No full-text available
To read the full-text of this research,
you can request a copy directly from the authors.
Abstract
In several neurological disorders including hyperhomocysteinemia, homocysteine (Hcy) accumulates in the brain, and acts as a potent neurotoxin. However, the molecular mechanisms induced by increased levels of Hcy in brain are not well understood. Here we show an activation of the extracellular signal-regulated kinases (ERK1 and ERK2) and the downstream nuclear targets Elk-1 and calcium/cAMP response element binding protein, in the hippocampus of cystathionine beta synthase deficient mice, a murine model of hyperhomocysteinemia. An ex vivo model of hippocampal slices allowed us to reproduce Hcy -induced ERK activation and to unravel the mechanisms responsible of this activation. Of interest, N-methyl-d-aspartate (NMDA), non-NMDA and metabotropic glutamate receptor antagonists all blocked Hcy -induced ERK activation. Moreover, the ERK activation was blocked in the presence of Na+-channel blocker tetrodotoxin, indicating the existence of a trans-synaptic activity in ERK activation by Hcy in hippocampal slices. The effects of Hcy on ERK cascade activation were also dependent on calcium influx, CaMK-II, PKC as well as PKA activation. Thus, altogether these data support a role of Hcy on ERK activation, via complex mechanisms, starting with a control of glutamate release, which in turn activates ionotropic and metabotropic receptor subtypes and produces increases in intracellular calcium levels.
... In addition, Hcy has been implicated to indirectly increase intracellular calcium activating ionotropic and metabotropic receptors [41], which, compared with glutamate, m the reasons to elicit different outcomes by such a combination of pharmacological actio [2,36,41]. ...
... In addition, Hcy has been implicated to indirectly increase intracellular calcium activating ionotropic and metabotropic receptors [41], which, compared with glutamate, m the reasons to elicit different outcomes by such a combination of pharmacological actio [2,36,41]. ...
... In addition, Hcy has been implicated to indirectly increase intracellular calcium levels by activating ionotropic and metabotropic receptors [41], which, compared with glutamate, may create the reasons to elicit different outcomes by such a combination of pharmacological actions of Hcy [2,36,41]. ...
Memory impairment has been shown to be associated with glutamate (Glu) excitotoxicity, homocysteine (Hcy) accumulation, and oxidative stress. We hypothesize that Glu and Hcy could damage neuronal cells, while astaxanthin (ATX) could be beneficial to alleviate the adverse effects. Using PC12 cell model, we showed that Glu and Hcy provoked a huge amount of reactive oxygen species (ROS) production, causing mitochondrial damage at EC50 20 and 10 mm, respectively. The mechanisms of action include: (1) increasing calcium influx; (2) producing ROS; (3) initiating lipid peroxidation; (4) causing imbalance of the Bcl-2/Bax homeostasis; and (5) activating cascade of caspases involving caspases 12 and 3. Conclusively, the damages caused by Glu and Hcy to PC12 cells can be alleviated by the potent antioxidant ATX.
... The data on homocysteine-induced activation of MAPK pathway confirm previous evidences from other Authors and ourselves in VSMC [11,[25][26][27], neurons [28], and other cell types [29,30]. ...
... This finding is in accordance with previous reports showing that NMDAr stimulation is deeply involved in the appearance of central nervous system symptoms in patients affected by hyperhomocysteinaemia [13,17,28]. Furthermore, studies which investigated homocysteine effects in neuronal cells of different areas of the central nervous system -such as hippocampus, cortex and striatum corpusproposed that homocysteine mediates MAPK activation likely by binding to glutamate site [13,17,28]. ...
... This finding is in accordance with previous reports showing that NMDAr stimulation is deeply involved in the appearance of central nervous system symptoms in patients affected by hyperhomocysteinaemia [13,17,28]. Furthermore, studies which investigated homocysteine effects in neuronal cells of different areas of the central nervous system -such as hippocampus, cortex and striatum corpusproposed that homocysteine mediates MAPK activation likely by binding to glutamate site [13,17,28]. ...
Introduction:
Exposure of vascular smooth muscle cells (VSMC) to homocysteine, at concentrations associated with an increased risk of cardiovascular events, enhances synthesis and secretion of Matrix Metalloproteinase-2 (MMP-2), which is involved in atherosclerotic plaque instabilization. This effect was prevented by inhibitors of Mitogen Activated Protein Kinase (MAPK) and Phosphatidylinositol 3-Kinase (PI3-K) pathways, allowing to hypothesize that homocysteine activates both these pathways, likely via a receptor-mediated mechanism. One possible receptor is N-methyl-D-aspartate receptor (NMDAr), which is expressed in VSMC and is involved in homocysteine effects in other cell types.
Materials and methods:
VSMC exposed to DL-homocysteine or NMDA (100 micromol/L for both; 5 min-8 hours), were investigated by measuring: i) phosphorylation of ERK1/2, p38MAPK (signaling molecules of MAPK pathway) and Akt and p70S6K (signaling molecules of PI3-K pathway) by western blot; ii) synthesis and secretion of MMP-2 (western blot); iii) activation of MMP-2 (gelatin zimography). To evaluate NMDAr involvement in the homocysteine effects, the experiments were repeated in the presence of a non-competitive NMDAr-antagonist MK-801 (50 micromol/L) or L-glycine (10 micromol/L), which inhibits NMDAr function by promoting its internalization.
Results:
DL-homocysteine and NMDA time-dependently increased: i) the phosphorylation of ERK1/2, p38 MAPK, Akt and p70S6K (ANOVA, p<0.0001); ii) the synthesis, secretion and activation of MMP-2. DL-homocysteine and NMDA effects were prevented by VSMC pre-incubation with MK-801 or high L-glycine concentrations.
Conclusions:
In human VSMC homocysteine-at concentrations associated with increased cardiovascular risk- activates MAPK and PI3-K pathways and MMP-2 synthesis and secretion through NMDA receptor, a potential mechanism involved in intracellular signaling in response to homocysteine in VSMC.
... It has also been hypothesized that Hcy can compete with GABA and directly activates group I metabotropic glutamate receptors, favoring substantial calcium influx [197]. HHcy can determine an extracellular signal-regulated kinase in the hippocampus, effect blocked by three types of glutamate receptor antagonists (NMDA, non-NMDA, and metabotropic receptors) [155,199]. By activating ionotropic and metabotropic receptors, HCY indirectly increases intracellular calcium level and activates several kinases [155,199]. ...
... HHcy can determine an extracellular signal-regulated kinase in the hippocampus, effect blocked by three types of glutamate receptor antagonists (NMDA, non-NMDA, and metabotropic receptors) [155,199]. By activating ionotropic and metabotropic receptors, HCY indirectly increases intracellular calcium level and activates several kinases [155,199]. It has also been evidenced that HHcy leads to a severe reduction of dopamine turnover in the striatum, and that probably occurs due to a high affinity for Hcy in the third loop of the D2 receptor, and therefore Hcy exerts an allosteric antagonist activity of D2 receptors [200,201]. ...
Homocysteine (Hcy) is a sulfur-containing amino acid that is generated during methionine metabolism. Hyperhomocysteinemia (HHcy) is typically defined as levels >15 micro mols/L. Elevated plasma levels of Hcy can be caused by the deficiency of either vitamin B12 or folate. The active role of homocysteine is quite ambivalent: many studies detected its potential impact on neurological events; others try to identify it as one of the possible risk factors of cardiovascular events, but with a complementary and secondary role. HHcy has been reported in many neurologic disorders, including cognitive impairment and stroke, independent of long-recognized factors such as hyperlipidemia, hypertension, diabetes mellitus, and smoking. Nowadays, homocysteine could be considered as a possible link between a common vascular risk factor and potential alterations in degenerative neuronal disorders. HHcy-induced oxidative stress, endothelial dysfunction, inflammation, smooth muscle cell proliferation, and endoplasmic reticulum stress; all these aspects have been considered to play an essential role in the pathogenesis of several diseases, including atherosclerosis, major stroke, and vascular dementia. Specific models of astrocytes impairment in HHcy-mice, which mimic small vessel disease, have been developed with a three-step investigation (at 6, 10, 14 weeks of B6, B9, and B12 detrimental diet in wild type HHcy mouse). These studies found out that after ten weeks on a diet (at the most after 14 weeks), end-feet disruption occurs. This phenomenon is concomitant to reduced vascular labeling for aquaporin -4-water channels, lower protein/mRNA levels for Kir4.1, and BK potassium channels, associated with a higher expression of MMP-9. The most exciting finding is that microglial activation in this mice model was evident since the precocious time of observation (6-week time) and precedes astrocytic changes. Our research aims to review the possible role of HHcy in neurodegenerative disease and small-vessel disease and to understand its pathogenic impact.
... Moreover, elevated hcy level has been associated with neuropsychiatric disorders such as schizophrenia, depression, bipolar disorder, and autism [4][5][6][7]. We showed that hhcys is accompanied by alterations in signaling pathways initiated by receptor tyrosine kinases (RTK), the ERK and Phosphatidylinositol 3-kinase (PI3K)/Akt pathways, and brain-derived neurotrophic factor/TrkB signaling pathway, in brain of CBS deficient mice, a murine model of hhcy [8][9][10][11][12]. ...
... Taken together, our It has been shown in primary cortical neurons that glutamate-induced triggering of the ionotropic N-methyl-D-aspartate (NMDA) receptor (NMDAR) was required for the enhanced and persistent Pi3k/ Akt-dependent NF-kappa B activation by the 75 kDa tumor necrosis factor receptor (TNFR2), indicating a positive cooperation of TNF and neurotransmitter-induced signal pathways [32]. We previously demonstrated that both mGluRs, NMDAR and non-NMDAR, cooperatively mediate hcy-induced ERK activation, indicating complex cellular mechanisms that render the hippocampus sensitive to hcy [10]. Expression of serine / threonine phosphatase PP2A, a phosphatase, that supress IkB phosphorylation by IKK, is inhibited by the PI3K/Akt pathway in primary cultures of neurons [33,34]. ...
Hyperhomocysteinemia due to cystathionine beta synthase (CBS) deficiency is associated with diverse cognitive dysfunction. Considering the role of the serine/threonine kinase DYRK1A, not only in developmental defects with life-long structural and functional consequences, but also in multiple neurodegenerative diseases, its protein expression and kinase activity has been analyzed in brain of heterozygous CBS deficient mice and found to be increased. We previously demonstrated that specific liver treatment with an adenovirus expressing Dyrk1A normalizes hepatic DYRK1A level and decreases hyperhomocysteinemia in mice with moderate to intermediate hyperhomocysteinemia. We here use a hepatocyte-specific recombinant adeno-associated viral (AAV) serotype 8-mediated DYRK1A gene therapy (AAV2/8-DYRK1A) to analyze the effect of hepatic Dyrk1A gene transfer on some altered molecular mechanisms in brain of mice with intermediate hyperhomocysteinemia. Our selective hepatic treatment alleviates altered DYRK1A protein level and signaling pathways in brain of mice, the MAPK/ERK and PI3K/Akt pathways initiated by receptor tyrosine kinase, the BDNF dependent TrkB pathway, and NFkB pathway. These results demonstrate the positive effect of AAV2/8-DYRK1A gene transfer on neuropathological and inflammatory processes in brain of mice with intermediate hyperhomocysteinemia.
... Through Hcy-NMDA binding, Hcy indirectly enhances calcium influx [132,135,136]. Glycine is the variable which conditions the Hcy effects [29]; when glycine is in normal concentration (10 umol/L), Hcy acts as a partial antagonist of the glycine site of the NMDA receptor, and it inhibits the receptor-mediated activity, acting as a neuroprotective factor [29,132]. ...
... Another study suggested that Hcy can activate the group I metabotrophic glutamate receptors [29,138]. This interaction can be demonstrated by blocking NMDA, non NMDA and metabotrophic glutamate receptor, via specific antagonists: a reduction of calcium influx can be observed, when binding these receptors [29,136]. ...
Vitamin B12 and folate are supplied via two major pathways, the conversion of homocysteine
to methionine and the conversion of methyl malonyl coenzyme A to succinyl coenzyme
A. Therefore, the defect in both the vitamins results in an increase in both serum homocysteine and
methylmalonic acid. Hence, homocysteine, vitamin B12, and folate are closely linked together in
the so-called one-carbon cycle, making vitamin B12 the necessary co-enzyme for the methyl
donation from 5-methyl-tetra-hydrofolate in tetra-hydro-folate, necessary for methionine
synthetase. Folate is a cofactor in one-carbon metabolism, and it promotes the remethylation of
homocysteine, which can cause DNA strand breakage, oxidative stress and apoptosis. Vitamin B12
and folate are involved in nucleic acid synthesis and in the methylation reactions, and their deficit
causes the inhibition of S-adenosylmethionine mediated methylation reactions, and through the
related toxic effects of homocysteine, a possible direct alteration of the vascular endothelium and
inhibition of N-methyl-D-Aspartate receptors take place. We discussed the possible and still
controversial role of homocysteine accumulation in cerebral pathologies, starting from vascular
events to neurodegeneration and to endothelium damage mechanism
... Several potential mechanisms linking high concentrations of homocysteine and cognitive impairment have been proposed. High tHcy levels have been shown to promote apoptosis in the hippocampus and to exert a direct neurotoxic effect, mediated by the activation of the Nmethyl-D-aspartate (NMDA) and non-NMDA glutamate receptors (Kruman et al., 2000;Poddar and Paul, 2009;Robert et al., 2005). Furthermore, high tHcy may affect cerebral arteries and arterioles by inducing an alteration of the cerebrovascular endothelium (Faraci and Lentz, 2004). ...
... There are a number of biological pathways via which the MTHFR C677T polymorphism may be related to cognitive impairment and dementia mainly mediated by high tHcy levels. Prolonged exposure to higher basal tHcy in MTHFR C677T TT carriers could exert a direct neurotoxic effect (Lipton et al., 1997;Robert et al., 2005). Furthermore, the MTHFR C677T mutation has been found to be associated with DNA hypomethylation (Castro et al., 2004). ...
... Studies examining the effects of homocysteine on neuronal cells have clearly demonstrated its neurotoxic effects. A variety of mechanisms of action have been proposed including but not limited to apoptosis, oxidative stress, over activation of glutamate receptors, mitochondrial dysfunctions and activation of caspases ( Ho et al., 2003;Kruman et al., 2000;Robert et al., 2005). Additionally, it has been shown that homocysteine induces activation of both N-methyl-D-aspartate (NMDA) and group I metabotropic glutamate receptors (mGluR), which may play a role in homocysteine mediated excitoxicity (Ho et al., 2002;Zieminska et al., 2003). ...
... Additionally, it has been shown that homocysteine induces activation of both N-methyl-D-aspartate (NMDA) and group I metabotropic glutamate receptors (mGluR), which may play a role in homocysteine mediated excitoxicity (Ho et al., 2002;Zieminska et al., 2003). Cytoplasmic calcium influx, as a consequence of both excitoxicity and oxidative stress has also been suggested to play a crucial role in homocysteine induced neurotoxicity ( Loureiro et al., 2008;Robert et al., 2005). Several studies have proposed that homocysteine induces neuronal apoptosis by damaging DNA: neuronal DNA strand breaks have been demonstrated in rat hippocampal neurons in response to elevated homocysteine ( Ho et al., 2002; Kruman et al., 2000Kruman et al., , 2002; Liu et al., 2009). ...
Although the neurotoxic effects of homocysteine have been well elucidated, the effects of homocysteine in astrocytes have received little attention until recently. Previously we have demonstrated that elevated levels of homocysteine caused significant metabolic changes and altered mitochondrial function in primary cultures of astrocytes. However, the mechanisms behind such alterations remain unclear. As homocysteine is a key metabolite in one-carbon metabolism the present study examined if the effects of homocysteine on astrocyte function are mediated through an epigenetic mechanism. Following exposure to homocysteine for 72h, global DNA methylation and H3K9 acetylation were examined using flow cytometric analysis. Total DNA methyltransferase activity and protein levels of DNA methyltransferase 3B were measured. Exposure to homocysteine resulted in global DNA hypomethylation (p<0.05) and histone hyperacetylation (p<0.05). Total DNA methyltransferase activity significantly decreased following exposure to homocysteine (from 11.5±3.9 to 6.0±1.7OD/h/mg protein, p<0.01) which was accompanied by a significant reduction in protein levels of DNA methyltransferase 3B (p<0.05). Treatment of astrocytes with the DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine, mimicked the functional changes induced by homocysteine. In conclusion, the results demonstrate significant epigenetic modifications following exposure to homocysteine in astrocytes and these changes seem to mediate functional alterations.
... Hcy trespasses blood-brain barrier through a specific carrier and therefore cerebrospinalfluid (CSF) Hcy concentrations are strictly correlated to its plasma levels (approximately 5-10%) [3]. The brain may be particularly vulnerable to high Hcy levels for two possible causes: 1) the two aforementioned major metabolic pathways for Hcy breakdown are missing in the brain [4]; 2) Hcy is directly responsible for several mechanisms inducing neurotoxicity [1] [5] [6] [7] [8]. ...
... Experimental studies indeed found that Hcy acts as an excitatory aminoacid by activating N-methyl-D-Aspartate (NMDA) and non-NMDA glutamate receptors [1] [5], induces mitochondrial dysfunction [6], free radicals [7] and cytosolic *Address correspondence to this author at the Department of Neurological Sciences, University of Bari, Ospedale Policlinico, Piazza Giulio Cesare 11, 70124, Bari, Italy; Tel: +390805592321; Fax: +390805478532; E-mail: lamberti@neurol.uniba.it calcium accumulation [8], and activates apoptotic pathways [1] [8]. ...
Homocysteine (Hcy) exerts multiple neurotoxic mechanisms that have been linked to the pathogenesis of neurodegenerative disorders. Several studies observed elevated plasma Hcy levels in Parkinson's Disease (PD) patients treated with L-dopa, compared to healthy controls and to patients with other neurodegenerative disorders.
We performed an overview of published evidences assessing the possible correlations between Hcy levels and the incidence or pathogenesis of PD.
A Medline literature search was performed to identify all available studies on Hcy and the incidence or pathophysiology of PD up to 30/09/2009.
30 studies were included in this overview (20 studies on humans, 10 experimental studies). The relationship between metilentetrahydrofolate-reductase genotype (the most common genetic cause of hyperhomocysteinemia) and the development of PD was contradictory. Dietary patterns and B-vitamins levels (important determinants of Hcy levels) were associated with a not-significant increased risk of PD in three prospective studies. Investigations on plasma and cerebrospinalfluid Hcy concentrations in L-dopa naive PD patients gave conflicting results; some studies observed increased Hcy levels in L-dopa naïve PD patients compared to controls, while others found no difference. In vitro, Hcy caused dose-dependent depletion of dopaminergic mesencephalic neurons, by numerous pathogenetic mechanisms. In vivo brain administration of Hcy induced motor and behavioural changes, similar to those observed in animal models of PD.
Based on the available data, the possibility that the hyperhomocysteinemia may contribute to the pathogenesis of PD remains uncertain. L-dopa treatment represents the major determinant of the hyperhomocysteinemia observed in PD.
... Excess Hcy can undergo auto-oxidation to generate hydrogen peroxide, superoxide and hydroxyl radicals to induce oxidative stress [88,89]. Elevated Hcy (homocysteinemia) can stimulate cytosolic calcium accumulation [90][91][92][93], mitochondrial dysfunction [94] and apoptotic pathway activation [91], which are pivotal factors in neurodegeneration. Increased levels of Hcy could also potentiate glutamate receptor activity via its oxidative metabolites HCSA and HCA, which are mixed excitatory agonists of NMDA and non-NMDA receptors [87,[95][96][97]. ...
Many pathogenetic mechanisms have been proposed for amyotrophic lateral sclerosis (ALS). Recently, there have been emerging suggestions of a possible role for the gut microbiota. Gut microbiota have a range of functions and could influence ALS by several mechanisms. Here, we review the possible role of gut-derived neurotoxins/excitotoxins. We review the evidence of gut symptoms and gut dysbiosis in ALS. We then examine a possible role for gut-derived toxins by reviewing the evidence that these molecules are toxic to the central nervous system, evidence of their association with ALS, the existence of biochemical pathways by which these molecules could be produced by the gut microbiota and existence of mechanisms of transport from the gut to the blood and brain. We then present evidence that there are increased levels of these toxins in the blood of some ALS patients. We review the effects of therapies that attempt to alter the gut microbiota or ameliorate the biochemical effects of gut toxins. It is possible that gut dysbiosis contributes to elevated levels of toxins and that these could potentially contribute to ALS pathogenesis, but more work is required.
... Hcy acts as a partial antagonist of the NMDA receptors [4,162,171,207,208], but when the glycine concentration is increased (like in the brain ischemia, in vasospasms, i.e., in prolonged migraine aura attack), even low doses of Hcy could act as an agonist of NMDA channels [212,213], inducing an enhancement of calcium flows [213]. HHCy promotes an extracellular signalregulated kinase activity in the hippocampus, regulated or blocked by three glutamate receptor antagonists (NMDA, not-NMDA, and metabotropic receptors) [154,214]. It has been suggested that Hcy could directly activate group I metabotropic glutamate receptors, favoring calcium influx currents [212]. ...
Homocysteine (Hcy) is a sulfur-containing amino acid generated during methionine metabolism, accumulation of which may be caused by genetic defects or the deficit of vitamin B12 and folate. A serum level greater than 15 micro-mols/L is defined as hyperhomocysteinemia (HHcy). Hcy has many roles, the most important being the active participation in the transmethylation reactions, fundamental for the brain. Many studies focused on the role of homocysteine accumulation in vascular or degenerative neurological diseases, but the results are still undefined. More is known in cardiovascular disease. HHcy is a determinant for the development and progression of inflammation, atherosclerotic plaque formation, endothelium, arteriolar damage, smooth muscle cell proliferation, and altered-oxidative stress response. Conversely, few studies focused on the relationship between HHcy and small vessel disease (SVD), despite the evidence that mice with HHcy showed a significant end-feet disruption of astrocytes with a diffuse SVD. A severe reduction of vascular aquaporin-4-water channels, lower levels of high-functioning potassium channels, and higher metalloproteinases are also observed. HHcy modulates the N-homocysteinylation process, promoting a pro-coagulative state and damage of the cellular protein integrity. This altered process could be directly involved in the altered endothelium activation, typical of SVD and protein quality, inhibiting the ubiquitin-proteasome system control. HHcy also promotes a constant enhancement of microglia activation, inducing the sustained pro-inflammatory status observed in SVD. This review article addresses the possible role of HHcy in small-vessel disease and understands its pathogenic impact.
... Hyperhomocysteinemia has been associated with several systemic conditions [7][8][9][10][11][12][13][14][15][16] . In particular, Hcy levels are significantly associated with high blood pressure 17 as they inhibit the synthesis of endothelial nitric oxide causing its dysfunction and damages the myocardium via the excessive production of reactive oxygen radicals 18,19 . ...
Background: Here, we assess the association between Hcy serum levels and periodontal status in a large representative sample of the National Health and Nutrition Examination Survey (NHANES).
Methods: We included 4,021 participants with a periodontal examination, medical self-reported data, BP and blood samples to determine complete blood count, C-reactive protein (CRP) and Hcy levels. We then calculated the periodontal inflamed surface area (PISA) and the periodontal epithelial surface area (PESA). Multivariable regression analysis explored the association between Hcy, periodontal measures and blood pressure. Mediation analysis was performed to understand the effect of age on the association of periodontitis with BP. Mediation analysis assessed the effect of PISA and PESA in the link between Hcy and BP.
Results: 4,021 participants fulfilled the inclusion criteria. Hcy levels showed significant correlations with systolic BP, diastolic BP, PISA, PESA and age. PESA showed to be significantly associated with Hcy both for the crude and adjusted models (p<0.01), but not PISA (p>0.05). In the association of Hcy with systolic BP, PISA significantly mediated 17.4% and PESA 0.9%. In the association of Hcy with diastolic BP, PISA significantly mediated 16.3% and PESA 47.2%.
Conclusions: This report shows that Hcy and periodontitis are associated. Also, both PISA and PESA significantly mediated the association of Hcy with systolic BP and diastolic BP. Future studies shall deepen the mechanisms by which homocysteine levels increase in a clinical situation of periodontitis.
... All those changes in vascular smooth muscle cells, provide further neurotoxic peculiarity to HHcy as a risk factor for neurodegenerative diseases. 111 Recently, an experimental model, linked the HHcy to increased oxidative stress, upregulated expression of proteins that promote blood coagulation, exacerbated blood-brain barrier dysfunction and promoted the infiltration of inflammatory cells into the cortex in traumatic brain injury (TBI). 112 ...
ABSTRACT: Homocysteine (Hcy) is a sulfhydryl-containing amino acid, and intermediate metabolite formed in metabolising methionine (Met) to cysteine (Cys); defective Met metabolism can increase Hcy. The effect of hyperhomocysteinemia (HHcy) on human health, is well described and associated with multiple clinical conditions. HHcy is considered to be an independent risk factor for common cardiovascular and central nervous disorders, where its role in folate metabolism and choline catabolism is fundamental in many metabolic pathways. HHcy induces inflammatory responses via increasing the pro-inflammatory cytokines and downregulation of anti-inflammatory cytokines which lead to Hcy-induced cell apoptosis. Conflicting evidence indicates that the development of the homocysteine-associated cerebrovascular disease may be prevented by the maintenance of normal Hcy levels. In this review, we discuss common conditions associated with HHcy and biochemical diagnostic workup that may help in reaching diagnosis at early stages. Furthermore, future systematic studies need to prove the exact pathophysiological mechanism of HHcy at the cellular level and the effect of Hcy lowering agents on disease courses.
... Through Hcy-NMDA binding, Hcy indirectly enhances calcium influx [69]. This is not a constant reaction, and it largely depends in the Glycine concentration; when glycine is in normal concentration (10 µmol/L), Hcy acts as a partial antagonist of the glycine site of the NMDA receptor, and it inhibits the receptor-mediated activity, acting as a neuroprotective factor [23,65]. ...
... This has been proposed as a mechanism in human subjects with HHcy caused by mutations in cystathionine β-synthase (CBS), the most common of the genetic causes of severe hyperhomocysteinemia, associated with increases up to 40-fold in fasting Hcy, and classic homocystinuria (congenital homocystinuria) or MTHFR genes [23] which has been associated with mild (13-24 µM) and moderate (25-60 µM) HHCy levels [9]. In neuronal degeneration and age-related neurodegenerative diseases, the Hcy neurotoxicity is due to troubled mechanisms of methylation and/or redox potentials triggering increased calcium influx [24], amyloid and tau protein accumulation [25,26], apoptosis, and neuronal death [27,28]. ...
Increased plasma homocysteine is a risk factor for several pathological disorders. The present review focused on the role of homocysteine (Hcy) in different population groups, especially in risk conditions (pregnancy, infancy, old age), and on its relevance as a marker or etiological factor of the diseases in these age groups, focusing on the nutritional treatment of elevated Hcy levels. In pregnancy, Hcy levels were investigated in relation to the increased risk of adverse pregnancy outcomes such as small size for gestational age at birth, preeclampsia, recurrent abortions, low birth weight, or intrauterine growth restriction. In pediatric populations, Hcy levels are important not only for cardiovascular disease, obesity, and renal disease, but the most interesting evidence concerns study of elevated levels of Hcy in autism spectrum disorder (ASD) and attention deficit hyperactivity disorder (ADHD). Finally, a focus on the principal pathologies of the elderly (cardiovascular and neurodegenerative disease, osteoporosis and physical function) is presented. The metabolism of Hcy is influenced by B vitamins, and Hcy-lowering vitamin treatments have been proposed. However, clinical trials have not reached a consensus about the effectiveness of vitamin supplementation on the reduction of Hcy levels and improvement of pathological condition, especially in elderly patients with overt pathologies, suggesting that other dietary and non-dietary factors are involved in high Hcy levels. The importance of novel experimental designs focusing on intra-individual variability as a complement to the typical case–control experimental designs and the study of interactions between different factors it should be emphasized.
... It has been proven that Hcy could be linked to neurodegeneration; Hcy (in tight relationship with higher glycine levels in the brain) is an agonist of the endogenous glutamate receptors, NMDA receptors [22], influencing calcium influx [16,23,24], as well as through a direct activation of the group I metabotropic glutamate receptors [25]. Much evidence suggests that Hcy potentiates the toxicity of Abeta 42 deposition [26][27][28] and increases amyloid's toxicity on the smooth vascular cells in the brain [29]. ...
... The neonatal resistance of spontaneous transmitter release to ROS appeared to result from immature mechanisms of intracellular Ca 2+ control (Shakirzyanova et al., 2016). The ability of HCY to increase intraterminal Ca 2+ (Loureiro et al., 2008;Robert et al., 2005) may underlie the higher sensitivity of spontaneous release to H2O2 in animals with hHCY. Moreover, HCY induced activation of protein kinase C (PKC) (Loureiro et al., 2008;Signorello et al., 2009) which restores the redox sensitivity of MEPPs in newborns (Shakirzyanova et al., 2016). ...
Enhanced levelsof homocysteine during pregnancy induce oxidative stress and contributeto many age-related diseases. In this study, we analyzed age-dependent synaptic modifications in developingneuromuscular synapsesof rats with prenatal hyperhomocysteinemia (hHCY). One of the main findings indicate that the intensity and the timing of transmitter release in synapses of neonatal (P6 and P10) hHCY rats acquired features of matured synaptictransmissionof adultrats. Theamplitude andfrequency of miniature end-plate currents (MEPCs) and evoked transmitter release werehigherin neonatal hHCY animalscompared to the control group. Analysis of the kineticsof neurotransmitter release demonstrated more synchronized release in neonatal rats with hHCY.At the same time lower release probability wasobserved in adults with hHCY. Spontaneous transmitter release in neonates with hHCY was inhibited byhydrogen peroxide (H2O2) whereas in controlsthis oxidant was effectiveonly in adultanimalsindicating ahigher susceptibility ofmotor nerve terminalsto oxidative stress. The morphology and theintensity of endocytosis of synaptic vesicles in motor nerve endingswas assessed using the fluorescence dye FM1-43. Adult-like synapses were found in neonates with hHCYwhich werecharacterized bya larger area of presynaptic terminals compared to controls.No difference in the intensity of FM1-43 fluorescence was observed between two groups of animals. Prenatal hHCY resulted in reducedmuscle strength assessed by the Paw Grip Endurancetest.Using biochemical assays we found an increased level of H2O2and lipid peroxidation products in the diaphragm muscles of hHCY rats. This was associated with a lowered activity of superoxide dismutase and glutathione peroxidase. Our data indicate that prenatalhHCY induces oxidative stress and apparent faster functional and morphological "maturation" of motor synapses.Ourresults uncover synaptic mechanisms of disrupted muscle function observed in hHCY conditions whichmay contribute tothe pathogenesis of motor neuronal diseases associated with enhanced level of homocysteine
LINK: https://authors.elsevier.com/c/1YrGK2dUamPJmk
... Quality assessment of each included studies was accessed and scored by two independent reviewers (Yun-Sheng Wang and Jun Ye) using the Newcastle-Ottawa Quality (NOS) Assessment Scale to determine the quality of selection of study participants, comparability, exposure and outcome. 13 Disagreements were discussed, and further adjudication was resolved by a third reviewer (Rong Zhang). ...
Objectives
Fibroblast growth factor-21 (FGF-21) plays an important role in glucose and lipid metabolism. This study aims to systemically review the evidence regarding the relationship between the FGF-21 levels and type 2 diabetes mellitus (T2DM), as well as the related influential factors.
Methods
Research related to plasma/serum FGF-21 levels in patients with T2DM and healthy controls were searched in PubMed, EMBASE and The Cochrane Library databases (up to 31 March 2017). Pooled standard mean difference (SMD) with 95% CI was calculated by fixed-effect or random-effect model analysis. Heterogeneity test was performed by the Q- statistic and quantified using I 2, and publication bias was evaluated using a funnel plot and Egger’s linear regression test.
Results
In total, 317 articles were obtained after searching databases, and 11 studies with 866 patients with T2DM and 629 controls were finally included. Meta-analysis revealed that, compared with the control group, the T2DM group had a significantly higher plasma/serum FGF-21 level (p < 0.001), with the SMD of 1.34% and 95% CI (0.70 to 1.98). Meta-regression analysis and subgroup analyses suggested that body mass index (BMI), triglycerides (TG) and total cholesterol (TC) were likely related to the observed FGF-21 differences between two groups.
Conclusions
Overall, our study suggests that patients with T2DM have significantly higher plasma/serum FGF-21 levels, and the FGF-21 levels were influenced by BMI, TC and TG.
... Through Hcy-NMDA binding, Hcy indirectly enhances calcium influx [69]. This is not a constant reaction, and it largely depends in the Glycine concentration; when glycine is in normal concentration (10 µmol/L), Hcy acts as a partial antagonist of the glycine site of the NMDA receptor, and it inhibits the receptor-mediated activity, acting as a neuroprotective factor [23,65]. ...
Homocysteine (Hcy) is a sulfur-containing amino acid that is generated during methionine metabolism. Physiologic Hcy levels are determined primarily by dietary intake and vitamin status. Elevated plasma levels of Hcy can be caused by deficiency of either vitamin B12 or folate. Hyperhomocysteinemia (HHcy) can be responsible of different systemic and neurological disease. Actually, HHcy has been considered as a risk factor for systemic atherosclerosis and cardiovascular disease (CVD) and HHcy has been reported in many neurologic disorders including cognitive impairment and stroke, independent of long-recognized factors such as hyperlipidemia, hypertension, diabetes mellitus, and smoking. HHcy is typically defined as levels >15 micromol/L. Treatment of hyperhomocysteinemia with folic acid and B vitamins seems to be effective in the prevention of the development of atherosclerosis, CVD, and strokes. However, data from literature show controversial results regarding the significance of homocysteine as a risk factor for CVD and stroke and whether patients should be routinely screened for homocysteine. HHcy-induced oxidative stress, endothelial dysfunction, inflammation, smooth muscle cell proliferation, and endoplasmic reticulum (ER) stress have been considered to play an important role in the pathogenesis of several diseases including atherosclerosis and stroke. The aim of our research is to review the possible role of HHcy in neurodegenerative disease and stroke and to understand its pathogenesis.
... Homocysteic acid, an oxidative product of homocysteine that is released in response to excitatory stimulation, acts an excitatory neurotransmitter by activating the NMDA receptor (Cuenod et al., 1990). Activation of both AMPA and NMDA receptors leads to increased intracellular calcium, which in turn leads to activation of several kinases (Robert et al., 2005). Overstimulation of these receptors due to HHcy can then lead to increased free radicals and caspases, which leads to apoptosis (Mattson and Shea, 2003) and neurodegeneration. ...
Behind only Alzheimer’s disease, vascular contributions to cognitive impairment and dementia (VCID) is the second most common cause of dementia, affecting roughly 10–40% of dementia patients. While there is no cure for VCID, several risk factors for VCID, such as diabetes, hypertension, and stroke, have been identified. Elevated plasma levels of homocysteine, termed hyperhomocysteinemia (HHcy), are a major, yet underrecognized, risk factor for VCID. B vitamin deficiency, which is the most common cause of HHcy, is common in the elderly. With B vitamin supplementation being a relatively safe and inexpensive therapeutic, the treatment of HHcy-induced VCID would seem straightforward; however, preclinical and clinical data shows it is not. Clinical trials using B vitamin supplementation have shown conflicting results about the benefits of lowering homocysteine and issues have arisen over proper study design within the trials. Studies using cell culture and animal models have proposed several mechanisms for homocysteine-induced cognitive decline, providing other targets for therapeutics. For this review, we will focus on HHcy as a risk factor for VCID, specifically, the different mechanisms proposed for homocysteine-induced cognitive decline and the clinical trials aimed at lowering plasma homocysteine.
... The quality of the studies was assessed and scored by two investigators independently (Ting-Yu Mu and Da-Hui Zhang) using the Newcastle-Ottawa quality assessment scale (NOS) for observational studies (Robert et al. 2005). In the case of disagreement, a third investigator (Cheng-Lu Zhang) was consulted. ...
Postpartum depression is a common complication of childbearing and up to 12 months postpartum. This study aimed to determine the prevalence of postpartum depressive mood (PDM) in China by performing a meta-analysis of published studies. Studies that reported the prevalence of PDM in China were identified by searching the PubMed, Embase, CNKI, and CQVIP databases. Three thousand, one hundred, and two articles were obtained, and after careful evaluation, 26 studies were finally included in the meta-analysis. The combined studies included a total of 7618 cases with 1621 cases of PDM. The studies were assessed on the basis of heterogeneity testing and the potential for publication bias. Stata software 11.0 was used to perform the meta-analysis. The random-effect model showed that the prevalence of PDM was 21% with a 95% confidence interval (CI) of 17–25%. PDM was the highest 0 to 1.5 months after delivery. PDM levels decreased to 10.4% (95% CI 9.7–11.1%, P < 0.001) after publication bias were corrected. Sensitivity analyses evaluated the stability of our results and showed no significant change when any single study was excluded. Subgroup analyses showed that region, instruments used, cut-off score, and time points for depression assessment were positively associated with PDM prevalence. The prevalence of PDM differed among regions, with South Central China and East China exhibiting the lowest prevalence. The prevalence was higher in regions with poor economic development, suggesting that more attention should be devoted to Southwest and North China and that the prevalence of PDM should be evaluated 0 to 1.5 months after delivery.
... They might affect neuronal plasticity (Streck et al., 2003), with different mechanisms in early or in adult life, and improve neurodegeneration , alter brain energy productions mechanisms (Streck et al., 2003), potentiate inflammation (Lazarewicz et al., 2003;Herrmann et al., 2006) and reduce the endothelium response to oxidation processes (Lazarewicz et al., 2003;Streck et al., 2003;Herrmann et al., 2006). Homocysteine regulates calcium inflow, via the activation of group I metabotropic glutamate receptors (Lipton et al., 1997;Lazarewicz et al., 2003;Robert et al., 2005;Herrmann et al., 2006;Obeid and Herrmann, 2006), and this has a relevance in the induction of brain lipid peroxidation process, and expanding the neural calcium-related apoptosis mechanism (Blom and Smulders, 2011). Moreover, homocysteine accumulation has an amyloidogenic effect by inducing the endoplasmic reticulum protein HERP, which potentiates the c-secretase activity and enhances the accumulation of AB1-40 in the brain (Mok et al., 2002;Seshadri et al., 2002). ...
Dementia is a worldwide health problem which affects millions of patients; Alzheimer’s
disease (AD) and subcortical vascular dementia (sVAD) are the two most frequent forms
of its presentation. As no definite therapeutic options have been discovered, different risk
factors for cognitive impairment have been searched for potential therapies. This report
focuses on the possible evidence that vitamin D deficiency and hyper-homocysteinemia
can be considered as two important factors for the development or the progression of
neurodegenerative or vascular pathologies. To this end, we assessed: the difference in
vascular risk factors and vitamin D-OH25 levels among groups of sVAD, AD, and healthy
age-matched controls; the association of folate, B12, homocysteine, and vitamin D with
sVAD/AD and whether a deficiency of vitamin D and an increment in homocysteine levels
may be related to neurodegenerative or vessel damages. The commonly-considered
vascular risk factors were collected in 543 patients and compared with those obtained
from a healthy old volunteer population. ANOVA group comparison showed that vitamin
D deficiency was present in demented cases, as well as low levels of folate and high
levels of homocysteine, more pronounced in sVAD cases. The statistical models we
employed, with regression models built, and adjustments for biochemical, demographic
and neuropsychiatric scores, confirmed the association between the three measures
(folate decrease, hyperhomocysteinemia and vitamin D decrease) and dementia, more
pronounced in sVAD than in AD.
... The quality of the studies was accessed and scored by two investigators independently (Peng Wang, Shi-Yang Guan) using the Newcastle-Ottawa quality assessment scale (NOS) for observational studies [10]. In the case of disagreement, a third investigator (Shu-Zhen Xu) was consulted. ...
This study aims to derive a more precise estimation on carotid intima-media thickness (CIMT) level in patients with rheumatoid arthritis (RA) and related factors. Studies published from January 1, 1982 to December 31, 2014 in English, which comparing CIMT between RA group and control group were searched in PubMed, Embase, and Cochrane Library databases. Heterogeneity test was performed, and publication bias was evaluated. Stata software 12.0 was used to perform the meta-analysis. Two-thousand one hundred sixty-three articles were obtained after searching databases, and 47 studies were finally included in the meta-analysis. The result of the analysis in random effect model showed that RA group had significantly higher CIMT than control group, with the standardized mean difference (SMD) of 1.04 and 95% CI (0.81,1.27). To evaluate the stability of our results, sensitivity analyses were performed, and the results showed no significant change when any one study was excluded. Subgroup analyses showed that region, race, age, BMI, and disease duration were associated with CIMT in RA patients. In summary, CIMT in RA patients is thicker than healthy controls, and it is influenced by region, race, age, BMI, and disease duration.
... The biological actions linking hyperhomocysteinemia to cognitive dysfunction remain unclear, but many studies have tried to establish relationships between hyperhomocysteinemia and altered signaling pathways (Robert et al. 2004(Robert et al. , 2005. Hcy is an intermediate in sulfur amino acid metabolism; once formed, Hcy may be metabolized by remethylation to methionine or transsulfuration to cysteine. ...
Hyperhomocysteinemia resulting from cystathionine beta synthase (CBS) deficiency can produce cognitive dysfunction. We recently found that CBS-deficient mice exhibit increased expression of the serine/threonine kinase dual-specificity tyrosine-(Y)-phosphorylation-regulated kinase 1A (DYRK1A) in the brain. When dysregulated, DYRK1A contributes to the neurodegeneration, neuronal death, and loss of function observed in neurodegenerative diseases. However, brain plasticity can be improved by interventions like enriched environment combined with voluntary exercise (EE/VE). The present study sought to assess the effects of EE/VE on molecular mechanisms linked to DYRK1A overexpression in the brain of CBS-deficient mice. EE/VE was applied to 3-month-old female CBS-deficient mice for 1 month. Without intervention, CBS-deficient mice exhibited increased DYRK1A and decreased brain-derived neurotrophic factor (BDNF) levels in the cortex and hippocampus. However, EE/VE rescued these altered DYRK1A and BDNF levels in the hippocampus of CBS-deficient mice. We conclude that exercise combined with enriched environment can restore the altered molecular mechanisms in the brain of CBS-deficient mice.
... This may be ascribed to decreased basal metabolism in catechol-O-methyltransferase (COMT)-mediated of endogenous dopamine due to dopamine depletion and alterations in methylation index by MPTP treatment272829. Experimental studies indicate that homocysteine acts as an excitatory amino-acid by activating NMDA receptors [30]. Thus, homocysteine induces mitochondrial dysfunction, free radicals and cytosolic calcium accumulation, and activation of apoptotic pathways [13,31,32]. ...
Background:
Modulation of neurogenesis that acts as an endogenous repair mechanism would have a significant impact on future therapeutic strategies for Parkinson's disease (PD). Several studies demonstrated dopaminergic modulation of neurogenesis in the subventricular zone (SVZ) of the adult brain. Levodopa, the gold standard therapy for PD, causes an increase in homocysteine levels that induces neuronal death via N-methyl-D-aspartate (NMDA) receptor. The present study investigated whether elevated homocysteine by levodopa treatment in a parkinsonian model would modulate neurogenesis via NMDA receptor signal cascade and compared the effect of levodopa and pramipexol (PPX) on neurogenic activity.
Methodology/principal findings:
Neurogenesis was assessed in vitro using neural progenitor cells (NPCs) isolated from the SVZ and in vivo with the BrdU-injected animal model of PD using 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. Modulation of homocysteine levels was evaluated using co-cultures of NPCs and astrocytes and PD animals. Immunochemical and Western blot analyses were used to measure neurogenesis and determine the cell death signaling. Levodopa treatment increased release of homocysteine on astrocytes culture media as well as in plasma and brain of PD animals. Increased homocysteine by levodopa led to increased apoptosis of NPCs through the NMDA receptor-dependent the extracellular signal-regulated kinase (ERK) signaling pathways. The administration of a NMDA antagonist significantly attenuated apoptotic cell death in levodopa-treated NPCs and markedly increased the number of BrdU-positive cells in the SVZ of levodopa-treated PD animals. Comparative analysis revealed that PPX treatment significantly increased the number of NPCs and BrdU-positive cells in the SVZ of PD animals compared to levodopa treatment. Our present study demonstrated that increased homocysteine by levodopa has a detrimental effect on neurogenesis through NMDA receptor-mediated ERK signaling pathway.
Conclusions/significance:
Modulation of levodopa-induced elevated homocysteine by NMDA antagonist or dopamine agonist has a clinical relevance for PD treatment in terms of adult neurogenesis.
... Cystathionine beta synthase (CBS), the first enzyme involved in the transsulfuration pathway, catalyzes the condensation of homocysteine with serine to form cystathionine. Homocysteine can also turn back to S-adenosylhomocysteine (SAH) via reversal of the SAH hydrolase (SAHH) reaction (Selhub 1999). Whereas the biological actions linking hyperhomocysteinemia to these neuronal abnormalities are not well understood , many studies tried to establish relationships between hyperhomocysteinemia and alterations of signalling pathways (Robert et al. 2004Robert et al. , 2005). Dyrk1A, which gene is localized on human chromosome 21, is a protein kinase that belongs to an evolutionarily conserved family of proteins known as DYRKs (dual-specificity tyrosine-(Y)-phosphorylation regulated kinase) that might be responsible for mental retardation in Down's syndrome patients (Ryu et al. 2010). ...
Hyperhomocysteinemia is associated with brain disease. However, biological actions linking hyperhomocysteinemia to neuronal abnormalities are not well understood. We recently found a relationship between Dyrk1A protein expression, a serine/threonine kinase that might be responsible for cognitive functions in Down's syndrome, and hepatic S-adenosylhomocysteine hydrolase (SAHH) activity, which plays a key role in S-adenosylmethionine-dependent methylation reactions. Considering the role of methylation and Dyrk1A in cognitive functions, the aim of this study was to investigate the relationship between Dyrk1A and SAHH activity in brain of hyperhomocysteinemic mice. We found an increase in Dyrk1A protein expression and activity in brain of hyperhomocysteinemic mice, concomitant with an increased SAHH activity. The effect of overexpression of protein Dyrk1A on SAHH activity was confirmed in brain of Dyrk1A transgenic mice, and additionally we found a positive correlation between Dyrk1A and SAHH activity. These observations suggest a potential effect of Dyrk1A on brain phenotypes linked to hyperhomocysteinemia.
... also been shown to be implicated in the pathogenesis of ALS (Kruman et al., 2000;Rowland and Shneider, 2001;Jara-prado et al., 2003;Robert et al., 2005;Obeid and Herrmann, 2006). A different line of evidence has shown that familial and SALS have similar pathological pathways (Al-Chalabi and Leigh, 2000). ...
Studies have revealed that elevated homocysteine levels can cause damage to motor neurons through multiple neurotoxic mechanisms, thus leading to the pathogenesis of amyotrophic lateral sclerosis (ALS). One way by which homocysteine levels are increased in the body is the consequence of methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms. Therefore, to address this question, we studied the MTHFR C677T and A1298C polymorphisms in 437 sporadic ALS (SALS) and 439 healthy controls to learn whether they were associated with SALS. The overall SALS were not associated with MTHFR C677T and A1298C polymorphisms (χ(2)=1.378; p=0.502; χ(2)=1.304; p=0.521, respectively). However, when we stratified results in terms of gender, we found that the MTHFR C677T polymorphism (χ(2)=6.376; p=0.041), T677T genotype (χ(2)=5.508; p=0.019; odds ratio [OR]=2.561; 95% confidence interval [CI]=1.142-5.744), C677C/A1298A (χ(2)=5.216; p=0.022; OR=0.424, 95% CI=0.199-0.900), and T677T/A1298A (χ(2)=6.639; p=0.010; OR=2.900; 95% CI=1.252-6.717) compound genotypes were associated with SALS in female patients only. Moreover, stratification of SALS according to the onset of disease indicated that there was no association between MTHFR C677T (χ(2)=1.565; p=0.457; A1298C χ(2)=3.461; p=0.177) polymorphisms and overall spinal onset SALS. Further stratification analysis according to gender revealed that there was a remarkable association between MTHFR C677T (χ(2)=9.728, p=0.008), T677T genotype (χ(2)=7.820; p=0.005; OR=3.126; 95% CI=1.361-7.178) and T allele (χ(2)=5.000; p=0.025; OR=1.711; 95% CI=1.067-2.745), and T677T/A1298A compound genotype (χ(2)=9.108; p=0.003; OR=3.540; 95% CI=1.494-8.387) and spinal onset female SALS only. Likewise, there was also association between MTHFR A1298C polymorphism (χ(2)=5.946; p=0.051) and the C1298C genotype (χ(2)=5.282; p=0.022; OR=2.524; 95% CI=1.125-5.658), and the C677T/C1298C compound genotype (χ(2)=7.155; p=0.007; OR=1.045; 95% CI=0.983-1.112) and bulbar onset SALS only in women. In conclusion, the evidence we provide here clearly shows that MTHFR C677T and A1298C polymorphisms are genetic risk factors for SALS in women in a gender-specific manner whether they are of spinal or bulbar onset.
... Previous studies have demonstrated that homocysteine enhances the activity of protein kinase A (PKA) and the phosphorylation of CREB in livers and hippocampal slices, respectively. 31,32 In this study, we observed that the levels of cAMP were markedly higher in quiescent livers of HHcy mice compared with those in quiescent livers of normal mice ( Figure 6B). In normal mice, a significant increase in hepatic cAMP levels occurred at 6 and 12 hours after PH ( Figure 6B). ...
Elevated homocysteine levels are defined as hyperhomocysteinemia (HHcy), a disorder that is associated with cardiovascular and neurodegenerative diseases as well as with hepatic fibrosis. Recent studies have shown that HHcy promotes hepatic injury by increasing oxidative stress. Although homocysteine induces cell cycle arrest in a variety of different cell types, it is not known whether HHcy has a definitive role in hepatocyte proliferation during liver regeneration. In this report, we investigated the effect of homocysteine on liver regeneration. Our results demonstrated that mice with HHcy exhibited an impairment in liver regeneration after partial hepatectomy, as measured by immunohistochemical staining of proliferation cell nuclear antigen and bromodeoxyuridine incorporation. Impaired proliferation was also correlated with reduced cyclin D1 induction and elevated expression levels of both p53 and p21Cip1. In addition, the phosphorylation of Akt, which plays an essential role in normal regeneration responses, was attenuated during the early phases of liver regeneration in HHcy mice. Our results also indicated that the cAMP/protein kinase A pathway mediated the inhibitory effect of homocysteine on liver regeneration. These findings provide evidence that impairment of liver regeneration by HHcy may result in delayed recovery from liver injury induced by homocysteine itself.
Elevated plasma level of homocysteine, also termed as hyperhomocysteinemia, is acknowledged as a significant and independent risk factor of Alzheimer's disease. However, the mechanistic insight has not been thoroughly elucidated yet. In this work, 3,5-dihydroxybenzyloxy was explored as the unique reaction trigger and integrated into the naphthalimide fluorophore via a carbamate linker to afford a new probe for •OH imaging. •OH treatment induced aromatic hydroxylation and subsequent elimination reaction to release the caged fluorophore, accompanied with a highly specific and sensitive turn-on fluorescence response. Cell imaging results revealed that excess homocysteine triggered overwhelming •OH production, which was mediated by N-methyl-d-aspartate receptor and NADPH oxidase, and the resultant •OH stress further initiated neuronal ferroptosis, also confirmed by western blot analyses. Additionally, hyperhomocysteinemic mouse models were established, and Alzheimer-like dementia of the mice was observed from behavioral tests. Most importantly, with this probe, cerebral •OH fluctuation was in situ visualized in live mice, which positively correlated with the severity of Alzheimer-like dementia induced by hyperhomocysteinemia. These results reveal that cerebral •OH stress may be the critical nexus linking hyperhomocysteinemia and Alzheimer's disease. This work provides a robust fluorescence probe for in situ visualizing the cerebral •OH fluctuations and illuminating critical insights into •OH contributions in brain disorders.
The cannabinoid system is ubiquitously present and is classically considered to engage in neural and immunity processes. Yet, the role of the cannabinoid system in the whole body and tissue metabolism via central and peripheral mechanisms is increasingly recognized. The present review provides insights in (i) how cannabinoid signaling is regulated via receptor-independent and -dependent mechanisms and (ii) how these signaling cascades (might) affect skeletal muscle plasticity and physiology. Receptor-independent mechanisms include endocannabinoid metabolism to eicosanoids and the regulation of ion channels. Alternatively, endocannabinoids can act as ligands for different classic (cannabinoid receptor 1 [CB1 ], CB2 ) and/or alternative (e.g., TRPV1, GPR55) cannabinoid receptors with a unique affinity, specificity, and intracellular signaling cascade (often tissue-specific). Antagonism of CB1 might hold clues to improve oxidative (mitochondrial) metabolism, insulin sensitivity, satellite cell growth, and muscle anabolism, whereas CB2 agonism might be a promising way to stimulate muscle metabolism and muscle cell growth. Besides, CB2 ameliorates muscle regeneration via macrophage polarization toward an anti-inflammatory phenotype, induction of MyoD and myogenin expression and antifibrotic mechanisms. Also TRPV1 and GPR55 contribute to the regulation of muscle growth and metabolism. Future studies should reveal how the cannabinoid system can be targeted to improve muscle quantity and/or quality in conditions such as ageing, disease, disuse, and metabolic dysregulation, taking into account challenges that are inherent to modulation of the cannabinoid system, such as central and peripheral side effects.
Objectives
This study was conducted to investigate the prevalence of vertebral fracture (VF) and its risk in patients with rheumatoid arthritis (RA) as compared to healthy individuals, and to explore the underlying risk factors.Methods
The electronic databases of PubMed, EMBASE, and the Cochrane Library were applied to search for the relevant literatures, which reported the prevalence of VF in both RA patients and healthy controls (up to Mar 31, 2021). The non-weighted prevalence of VF, pooled estimates of odds ratio (OR), and its 95% confidence intervals (CI) were calculated with the use of random-effects model; between-study heterogeneity was evaluated by Cochrane Q statistic, then was quantified with I2. Publication bias was evaluated using Egger’s linear regression test.ResultsA number of 867 literatures were identified after searching for online databases, based on the inclusion and exclusion criteria, 11 eligible studies, which comprising 3805 RA patients and 59,517 healthy participants, were finally incorporated in meta-analysis. The results showed that RA patients had an increased prevalence of VF (20.29 vs 8.63%), and an elevated risk for VF (OR = 3.04, 95% CI 1.97–4.71) as compared to healthy population. Additional subgroup analysis suggested that age, body mass index (BMI), disease activity, and drug therapy might be associated with risk of VF in RA.Conclusions
Overall, our study demonstrated an increased risk of VF in patients with RA, suggesting that age, race, BMI, disease activity, and drug therapy may be represented as risk factors contributing to the occurrence of VF in RA.
Key Points
• RA patients had the increased prevalence and risk of vertebral fracture (VF) as compared to healthy population.
• Age, race, BMI, disease activity, and drug therapy might be associated with VF in RA.
• Our findings would be helpful for the early evaluation of RA patients with high VF risk.
Background and aim:
The relationship between the Helicobacter pylori (H. pylori) infection and homocysteine is unclear. We evaluated the effect of H. pylori on serum homocysteine in a healthy Chinese population.
Methods:
A total of 21,184 individuals aged over 18 years underwent 13 C/14 C urease breath test (13 C/14 C-UBT) and blood tests, and 5042 individuals with follow-up intervals greater than 6 months. Homocysteine levels are classified according to the Chinese expert consensus.
Results:
The rates of H. pylori infection of normal level, mild level, moderate level, and severe level were 40.9%, 43.8%, 45.8% and 46.6% respectively (P = 0.000). H. pylori infection increased the risk of higher homocysteine concentration (OR = 1.406, P = 0.000). In the case-control study, the rates of persistent negative, new infection, persistent infection, and eradication infection was 43.6%, 11.2%, 22.9% and 22.3% respectively. The percentage of changes in serum homocysteine levels varied significantly among the different H. pylori infection statuses only in mild level (P = 0.024). Mean changed homocysteine values were higher in the subgroup of persistent infection than in the persistent negative subgroup (P = 0.004) and the eradication infection subgroup (P = 0.034). Serum homocysteine values were elevated only in the subgroup with over three years' interval time and persistent infection (n = 107, mean paired differences = 1.1±4.6 umol/L, P = 0.014).
Conclusions:
There is a relationship between H. pylori and serum homocysteine, and persistent infection leads to elevation of the latter.
This study explored the potential effects of mild hyperhomocysteinemia (HHcy) on the blood–brain barrier (BBB) and neuroinflammation. Seven-week-old male wild-type C57BL/6 mice were fed normal mouse chow (the control group) or a methionine-enriched diet (the HHcy group) for 14 weeks. Mice in the HHcy group exhibited a slight increase in serum Hcy levels (13.56 ± 0.61 μmol/L). Activation of the ERK signaling pathway, up-regulation of matrix metalloproteinase-9 (MMP-9), and degradation of tight junction proteins (occludin and claudin-5) were observed in both the cerebral cortex and hippocampus of mice with mild HHcy. However, microglia were not activated and the levels of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) were not changed in either the cerebral cortex or hippocampus of mice with mild HHcy. Moreover, the signaling activity of STAT3 also did not differ significantly between the two groups. These findings demonstrate that the BBB is highly vulnerable to homocysteine insult. Even a slight increase in serum homocysteine levels up-regulates MMP-9 expression and disrupts the BBB integrity. Meanwhile, microglia activation or the STAT3 pathway might not contribute to the effects of mild HHcy on the brain.
Backgrounds:
Rheumatoid arthritis (RA) is a systemic autoimmune disease that confers one of the strongest risks for cardiovascular disease (CVD) morbidity and mortality than in general population. Pulse wave velocity (PWV) and augmentation index (AIx) are composite measures of arterial stiffness (AS) and associated with CV risk.
Aim of the study:
The aim of this study was to systemically review the evidence regarding the relationship between PWV, AIx and RA, as well as underlying influential factors.
Methods:
Eligible literatures were searched in PubMed, EMBASE and The Cochrane Library published up to February 28, 2019 in English. The pooled weight mean difference (WMD) with its 95% confidence interval (CI) was calculated using random-effect model analysis.
Results:
A total of 38 studies were finally incorporated in the meta-analysis. The results indicated that, compared to controls, RA patients had significantly increased levels of carotid-femoral (cf)-PWV (WMD = 1.10 m/s, 95% CI: 0.84-1.35), brachial-ankle (ba)-PWV (WMD = 0.20 m/s, 95% CI: 0.12-0.28), cartoid-radial (cr)-PWV (WMD = 0.51 m/s, 95% CI: 0.23-0.79), AIx (WMD = 4.79%, 95% CI: 1.34-8.24) and AIx normalized to a 75 beats/minute heart rate (AIx@75) (WMD = 5.78%, 95% CI: 3.82-7.74) (all p <0.001). Meta-regression and subgroup analysis demonstrated significant association of cf-PWV with age, disease duration and erythrocyte sedimentation rate (ESR) in RA.
Conclusions:
Overall, there is increased PWV level in patients with RA, and this alteration is associated with age, disease duration and ESR.
Arterial stiffness (AS) is a predictor of coronary artery outcomes in patients with cardiovascular disease (CVD). Carotid–femoral pulse wave velocity (cf-PWV) is a commonly used method for assessing AS. This study aimed to assess the relationship between cf-PWV and clinical CVD events. Of the 786 studies identified, 19 studies were included in the final meta-analysis. Meta-analysis revealed that participants with high cf-PWV by 1 standard deviation (SD), 1 m/s, and cutoff points have a high pooled relative risk for CVD events (1 SD: 1.25, 95% confidence interval [CI]: 1.19-1.31; 1 m/s: 1.12, 95% CI: 1.07-1.18; and cutoff points: 1.80, 95% CI: 1.45-2.14) and CVD mortality (1 SD: 1.23, 95% CI: 1.15-1.31; 1 m/s: 1.09, 95% CI: 1.04-1.14; and cutoff points: 1.85, 95% CI: 1.46-2.24). In addition, we found that the predictive value of increased AS was higher in patients with higher disease risk for total CVD events and CVD mortality than in other patients. Carotid–femoral pulse wave velocity is a useful biomarker to improve the prediction of CV risk for patients and identify high-risk populations who may benefit from aggressive CV risk factor management.
Systemic lupus erythematosus (SLE) is associated with increased cardiovascular risk. This study aims to derive a more precise estimation on pulse wave velocity (PWV) level in patients with SLE and related factors. A literature search was performed using PubMed, EMBASE, and The Cochrane Library databases, studies published up to February 28, 2017, in English. Pooled standardized mean difference (SMD) with 95% confidence interval (CI) was calculated by fixed-effect or random-effect model analysis. Heterogeneity test was performed by the Q statistic and quantified using I²; publication bias was evaluated using a funnel plot and Egger’s linear regression test. Of 156 studies found, 27 met eligibility criteria, and 14 studies were finally included in the meta-analysis. Meta-analysis revealed that the SLE group had significantly higher PWV levels than the control group; SMD = 0.56 and 95% CI (0.30-0.82). Subgroup analyses showed that body mass index (BMI), sample size, and disease duration were associated with PWV in patients with SLE. Overall, our study suggests that patients with SLE have a higher PWV level, and it is associated with BMI, sample size, and disease duration.
Background and purpose:
Vascular inflammation, including the expression of inflammatory cytokines in endothelial cells, plays a critical role in hyperhomocysteinemia-associated vascular diseases. Cathepsin V, specifically expressed in humans, is involved in vascular diseases through its elastolytic and collagenolytic activities. The aim of this study was to determine the effects of cathepsin V on the L-homocysteine (L-Hcy)-induced vascular inflammation.
Experimental approach:
A high methionine diet-induced hyperhomocysteinemic mice model was used to assess the cathepsin V expression and vascular inflammation. Cultures of HUVECs were challenged with L-Hcy and cathepsin L/V inhibitor SID to assess the pro-inflammatory effects of cathepsin V. Transfection and anti-sense techniques were utilized to investigate the influences of cathepsin V on the dual specificity protein phosphotases (DUSPs) and mitogen-activated protein kinase (MAPK) pathways.
Key results:
We revealed that cathepsin L (human cathepsin V homologous) was increased in the thoracic aorta endothelial cells of hyperhomocysteinemic mice, L-Hcy promoted cathepsin V expression in HUVECs. SID suppressed the activity of cathepsin V, reversed up-regulation of inflammatory cytokine (IL-6, IL-8, TNFα), adhesion and chemotaxis of leukocyte, and vascular inflammation induced by L-Hcy in vivo and in vitro. Increased cathepsin V promotes the degradation of DUSP6 and DUSP7, phosphorylation and subsequent nuclear translocation of ERK1/2 , phosphorylation of STAT1 and expression of IL-6, IL-8, TNFα.
Conclusion and implications:
This study has therefore delineated a novel mechanism that L-Hcy-induced cathepsin V mediated vascular endothelial inflammation under high homocysteine condition partly via ERK1/2 /STAT1 pathway. This mechanism could represent a potential therapeutic target in hyperhomocysteinemia-associated vascular diseases.
Objectives
To derive a more precise comparison of flow-mediated dilatation (FMD%) of the brachial artery between patients with rheumatoid arthritis (RA) and normal controls by performing a meta-analysis of appropriate studies.
Methods
PubMed and EMBASE databases were searched for all relevant articles. STATA (V.12.0) software was used to perform the meta-analysis. Quality estimation of all appropriate studies was evaluated according to the Newcastle-Ottawa Scale (NOS). Standardised mean difference (SMD) with 95% CIs were calculated with a random-effects model. The Cochrane Q test and I² statistic were used to evaluate the heterogeneity. Funnel plot and Egger's test were conducted to assess the publication bias.
Results
In total, 464 articles were obtained after searching the two databases. Ten studies were included in the meta-analysis on the basis of the inclusion and exclusion criteria. Significant heterogeneity was observed among these 10 studies (Q=102.89, p<0.001, I²=91.3%) with random-effects modelling. The results showed that the RA group had significantly lower FMD% (SMD: −1.405; 95% CI −1.992 to −0.817; p<0.001) than the control group. Egger's test (p=0.004) indicated that the funnel plot showed a skewed or asymmetrical shape and publication bias existed. Sensitivity analyses suggested the robustness and credibility of our results.
Conclusions
FMD% in patients with RA is significantly decreased compared with healthy controls. FMD% is an important early marker of atherosclerosis. It may be used as a parameter to forecast cardiovascular disease in patients with RA.
Liver diseases lead to a complex syndrome characterized by neurological, neuro-psychiatric and motor complications, called hepatic encephalopathy, which is prevalent in patients and animal models of acute, sub-chronic and chronic liver failure. Although alterations in GABAergic, glutamatergic, cholinergic and serotonergic neuronal functions have been implicated in HE, the molecular mechanisms that lead to HE in chronic liver disease (CLD) is least illustrated. Due to hepatocellular failure, levels of ammonia and homocysteine (Hcy), in addition to others, are found to increase in the brain as well as plasma. Hcy, a non-protein forming amino acid and an excitotoxin, activates ionotropic glutamate (n-methyl-d-aspartate; NMDA) receptors, and thereby leads to influx of Ca(2+) into neurons, which in turn activates several pathways that trigger oxidative stress, inflammation and apoptosis, collectively called excitotoxicity. Elevated levels of Hcy in the plasma and brain, a condition called Hyperhomocysteinemia (HHcy), and the resultant NMDA receptor-mediated excitotoxicity has been implicated in several diseases, including Parkinson's disease and Alzheimer's disease. Although, hyperammonemia has been shown to cause excitotoxicity, the role of HHcy in the development of behavioral and neurochemical alterations that occur in HE has not been illustrated yet. It is hypothesized that CLD-induced HHcy plays a major role in the development of HE through activation of NMDA receptors. It is further hypothesized that HHcy synergizes with hyperammonemia to activate NMDA receptor in the brain, and thereby cause oxidative stress, inflammation and apoptosis, and neuronal loss that leads to HE.
Copyright © 2015. Published by Elsevier Ltd.
Hyperhomocysteinemia is a risk factor for neurological diseases, but the underlying pathophysiology has not been adequately explained. Mild hyperhomocysteinemia, which is sometimes associated with a low plasma level of vitamin B9, B12 and folic acid, is responsible in the toxicity in neural cell by activating NMDA receptor. Indeed, even if vitamin supplementation has clearly proven its efficiency on lowering plasma levels of homocysteine, recent studies do not show any positive effect of vitamin therapy on cognitive function. The hypothesis that this therapy is inefficient has been recently reinforced by two randomized trials on the effects of vitamin supplementation. Several hypotheses still need to be explored: Mechanisms of homocysteine toxicity and that of total uselessness of vitamin supplementation; the possible need to complete the actual data with further, more powerful studies in order to prove the role of homocysteine in the development of neurodegenerative diseases and a clinical effect of vitamin therapy.
Hyperhomocysteinemia due to cystathionine beta synthase (CBS) deficiency is associated with diverse brain disease. Whereas the biological actions linking hyperhomocysteinemia to the cognitive dysfunction are not well understood, we tried to establish relationships between hyperhomocysteinemia and alterations of signaling pathways. In the brain of CBS-deficient mice, a murine model of hyperhomocysteinemia, we previously found an activation of extracellular signal-regulated kinase (ERK) pathway and an increase of Dyrk1A, a serine/threonine kinase involved in diverse functions ranging from development and growth to apoptosis. We then investigated the relationship between Dyrk1A and the signaling pathways initiated by receptor tyrosine kinases (RTK), the ERK and PI3K/Akt pathways. We found a significant increase of phospho-ERK, phospho-MEK, and phospho-Akt in the brain of CBS-deficient and Dyrk1a-overexpressing mice. This increase was abolished when CBS-deficient and Dyrk1A-transgenic mice were treated with harmine, an inhibitor of Dyrk1A kinase activity, which emphasizes the role of Dyrk1A activity on ERK and Akt activation. Sprouty 2 protein level, a negative feedback loop modulator that limits the intensity and duration of RTK activation, is decreased in the brain of CBS-deficient mice, but not in the brain of Dyrk1A transgenic mice. Furthermore, a reduced Dyrk1A and Grb2 binding on sprouty 2 and an increased interaction of Dyrk1A with Grb2 were found in the brain of Dyrk1A transgenic mice. The consequence of Dyrk1A overexpression on RTK activation seems to be a decreased interaction of sprouty 2/Grb2. These observations demonstrate ERK and Akt activation induced by Dyrk1A in the brain of hyperhomocysteinemic mice and open new perspectives to understand the basis of the cognitive defects in hyperhomocysteinemia.
Hyperhomocysteinemia is a risk factor for neurological diseases, but the underlying pathophysiology has not been adequately explained. Mild hyperhomocysteinemia, which is sometimes associated with a low plasma level of vitamin B9, B12 and folic acid, is responsible in the toxicity in neural cell by activating NMDA receptor. Indeed, even if vitamin supplementation has clearly proven its efficiency on lowering plasma levels of homocysteine, recent studies do not show any positive effect of vitamin therapy on cognitive function. The hypothesis that this therapy is inefficient has been recently reinforced by two randomized trials on the effects of vitamin supplementation. Several hypotheses still need to be explored: Mechanisms of homocysteine toxicity and that of total uselessness of vitamin supplementation; the possible need to complete the actual data with further, more powerful studies in order to prove the role of homocysteine in the development of neurodegenerative diseases and a clinical effect of vitamin therapy.
Homocysteine (HCY) is a known risk factor for neuronal diseases. We here report that HCY (10–1000 μM) interfered bi-directionally with long-term potentiation (LTP) in hippocampal slices, causing an impairment at concentrations <100 μM, and enhancement ≥500 μM. By comparison, NMDA unidirectionally reduced LTP, whereas l-cysteine led to facilitated LTP. Such HCY-induced alterations in neuronal communication may contribute to cognitive failure in dementia.
The study determines whether hyperhomocysteinemia (risk factor of brain ischemia) alone or in combination with ischemic preconditioning (IPC) affects the ischemia-induced changes in gene expression of secretory pathways Ca(2+)-ATPase (SPCA1). Hyperhomocysteinemia was induced by subcutaneous administration of homocysteine (Hcy; 0.45 µmol/g body weight) twice a day at 8 h intervals for 14 days. Rats were preconditioned by 5 min ischemia and 2 days later, 15 min of global forebrain ischemia was induced by four vessel occlusion. We observed that hyperhomocysteinemia significantly decreased the level of SPCA1 mRNA in the cortex. Pre-ischemic challenge was noticeable in both brain areas. In the cortex, pre-ischemia in Hcy group led to the abrupt stimulation of the mRNA expression by 249% within the Hcy ischemic group and by 321% in the Hcy control. Values further exceeded those observed in the naive control. In the hippocampus, the differences between naive and Hcy groups were not observed. IPC initiated elevation of mRNA expression to 159% (p < 0.05) of control with Hcy and to 131% (p < 0.01) of ischemia with Hcy, respectively. Documented response of SPCA gene to IPC in hyperhomocysteinemic group might suggest a correlation of SPCA expression consistent with the role of cross-talks between intracellular Ca(2+) stores including secretory pathways in the tolerance phenomenon.
Folate and vitamin B12 are involved in homocysteine metabolism and are critical to the methylation of DNA. We aimed to assess plasma vitamin B12 (pB12), plasma folate (pFol), and red cell folate (rcFol) in women and their infants during pregnancy and after birth. Maternal biomarkers were tested as predictors of infant biomarkers, including plasma homocysteine (pHcy), at age 6 months. Participants (n = 153) were recruited at the John Hunter Hospital, Australia. Maternal fasting blood samples were collected at 20 and 36 weeks gestation, and at 14 and 27 weeks postpartum. Fifty healthy, term infants provided non-fasting samples at age 6 months. Plasma homocysteine data were available for 16 infants at age 6 months. Maternal pB12 concentrations fell by 16% from 20 to 36 weeks gestation, but had recovered by 14 weeks postpartum. Maternal rcFol concentrations fell by 31% from 20 weeks gestation to 27 weeks postpartum. Infants breastfed at 6 months had lower pB12 (median 159 vs. 402 pmol/L, n = 23 vs. 18, P < 0.01) and folate (median folate z-score -0.58 vs. 0.85, n = 23 vs. 17, P < 0.01), and higher pHcy (median 11.9 vs. 7.3 μmol/L, n = 8 vs. 6, P < 0.01), than those on infant formula. Maternal pregnancy pFol, but not pB12, inversely predicted infant pHcy, after adjustment for the infant's current pB12 (P = 0.04). Changes in maternal B12 and folate occur during pregnancy and after birth. Infant homocysteine metabolism may be regulated through maternal folate concentrations during pregnancy and postnatal feeding.
Both elevated homocysteine and decreased hydrogen sulfide (H(2)S) are observed in the brains of Alzheimer's disease (AD) patients. Reactive oxygen species (ROS) overproduction contributes to the neurotoxicity of homocysteine; however, H(2)S is an endogenous antioxidant gas. Therefore, the aim of this study was to investigate whether the imbalance of proportion to this endogenous protective antioxidant gas is involved in homocysteine-caused neurotoxicity. We show that homocysteine inhibits the generation of endogenous H(2)S and the expression and activity of cystathionine-β-synthetase (CBS), the main enzyme responsible for the generation of H(2)S in PC12 cells. S-Adenosylmethionine, an activator of CBS, not only prevents homocysteine-induced inhibition of endogenous H(2)S production but also attenuates homocysteine-triggered cytotoxicity and accumulation of ROS. We find that activation of ERK1/2 occurs in homocysteine-treated PC12 cells and blockade of ERK1/2 with U0126 abolished the homocysteine-induced cytotoxicity and inhibitory effect on endogenous H(2)S generation. These results indicate that homocysteine neurotoxicity involves reduction of H(2)S production, which is caused by inhibition of CBS and mediated by activation of ERK1/2. Our study suggests a promising future of H(2)S-based therapies for neurodegenerative diseases such as AD.
Hyperhomocysteinaemia (HHcy) has been identified as a cardiovascular risk factor for neurodegenerative brain diseases. The aim of the present study was to investigate the effects of short (5 months) or long (15 months) HHcy in Sprague–Dawley rats in vivo. Short- and long-term HHcy differentially affected spatial memory as tested in a partially baited eight-arm radial maze. HHcy significantly reduced the number of choline acetyltransferase (ChAT)-positive neurons in the basal nucleus of Meynert and ChAT-positive axons in the cortex only after short-term but not long-term treatment, while acetylcholine levels in the cortex were decreased at both time points. Nerve growth factor (NGF) was significantly enhanced in the cortex only after 15 months of HHcy. HHcy did not affect cortical levels of amyloid precursor protein, beta-amyloid(1-42), tau and phospho-tau181 and several inflammatory markers, as well as vascular RECA-1 and laminin density. However, HHcy induced cortical microbleedings, as illustrated by intensive anti-rat IgG-positive spots in the cortex. In order to study the regulation of the key enzyme ChAT, organotypic rat brain slices were incubated with homocysteine, which induced a decline of ChAT that was counteracted by NGF treatment. In conclusion, our data demonstrate that chronic short- and long-term HHcy differentially caused memory impairment, cholinergic dysfunction, NGF expression and vascular microbleedings.
Hyperhomocysteinemia has been associated with cognitive dysfunction and dementia. The incidence of dementia in Parkinson's Disease (PD) patients is higher than in the general population and plasma Homocysteine concentrations are increased in L-dopa treated PD patients.
We evaluated the possible correlations between L-Dopa related hyperhomocysteinemia and cognitive dysfunction in PD.
A Medline literature search was performed to identify all published studies on Homocysteine and cognitive dysfunction and dementia during the course of PD from 1966 to 31/03/2010.
Sixteen studies were found for review; ten studies focused on homocysteine and cognitive dysfunction in PD patients, five on homocysteine and PD dementia and two on homocysteine and markers of neurodegeneration in PD. The design of the study was retrospective in 14 studies, while 2 had a prospective design, with a variable follow-up period (from 24-weeks to 2 years). In most of the studies plasma homocysteine levels significantly correlated with cognitive functions, dementia and markers of neurodegeneration in PD patients. However, some studies did not confirm these findings. Several factors may concur to explain these partially conflicting results, including the retrospective design of the studies, their small sample size, their high percentage of excluded patients, and the use of a wide range of neuropsychological tasks in assessment of cognitive dysfunctions across the available studies.
Available data seem to indicate a potential role of L-dopa related hyperhomocysteinemia on cognitive impairment and dementia during the course of PD.
Hyperhomocysteinemia has been implicated in dementia and neurodegenerative disease. Physiological homocysteine concentrations did not result in apoptosis in SH-SY5Y cells in the present study. The apoptosis was recognized in millimolar level of homocysteine. However, SH-SY5Y cell death was observed following exposure to micromolar level of homocysteine in combination with copper. Exposure to 250microM homocysteine and 10microM CuCl(2) for one day decreased cell viability by 40%. Homocysteine and copper caused apoptosis, because hallmarks of apoptosis were recognized, such as loss of mitochondrial membrane potential, TUNEL-positive cells, release of cytochrome c from mitochondria, and caspase-3 activation, but not nucleosomal DNA fragmentation. Homocysteine and copper generated the intracellular reactive oxygen species, and homocysteine and copper-induced apoptosis was due to an accumulation of intracellular reactive oxygen species, which was inhibited by catalase. Pan-caspase inhibitor, z-VAD-fmk, could not completely inhibited homocysteine and copper-induced cell death. Homocysteine and copper also caused the nuclear translocation of apoptosis-inducing factor. These results suggested that homocysteine and copper induced not only caspase-dependent apoptosis but also caspase-independent apoptosis-inducing factor related apoptosis.
The PSD-95/SAP90 family of proteins has recently been implicated in the organization of synaptic structure. Here, we describe the isolation of a novel Ras-GTPase activating protein, SynGAP, that interacts with the PDZ domains of PSD-95 and SAP102 in vitro and in vivo. SynGAP is selectively expressed in brain and is highly enriched at excitatory synapses, where it is present in a large macromolecular complex with PSD-95 and the NMDA receptor. SynGAP stimulates the GTPase activity of Ras, suggesting that it negatively regulates Ras activity at excitatory synapses. Ras signaling at the postsynaptic membrane may be involved in the modulation of excitatory synaptic transmission by NMDA receptors and neurotrophins. These results indicate that SynGAP may play an important role in the modulation of synaptic plasticity.
Studies by various investigators have indicated that elevated levels of plasma homocyst(e)ine are strongly associated with the occurrence of occlusive vascular diseases. With the eventual aim of determining whether or not elevated plasma homocyst(e)ine concentrations are directly causative of cardiovascular diseases, we have generated mice that are moderately and severely homocyst(e)inemic. Homologous recombination in mouse embryonic stem cells was used to inactivate the cystathionine beta-synthase [L-serine hydrolyase (adding homocysteine), EC 4.2.1.22] gene. Homozygous mutants completely lacking cystathionine beta-synthase were born at the expected frequency from matings of heterozygotes, but they suffered from severe growth retardation and a majority of them died within 5 weeks after birth. Histological examination showed that the hepatocytes of homozygotes were enlarged, multinucleated, and filled with microvesicular lipid droplets. Plasma homocyst(e)ine levels of the homozygotes were approximately 40 times normal. These mice, therefore, represent a model for severe homocyst(e)inemia resulting from the complete lack of cystathionine beta-synthase. Heterozygous mutants have approximately 50% reduction in cystathionine beta-synthase mRNA and enzyme activity in the liver and have twice normal plasma homocyst(e)ine levels. Thus, the heterozygous mutants are promising for studying the in vivo role of elevated levels of homocyst(e)ine in the etiology of cardiovascular diseases.
To understand how extracellular signals may produce long-term effects in neural cells, we have analyzed the mechanism by which neurotransmitters and growth factors induce phosphorylation of the transcription factor cAMP response element binding protein (CREB) in cortical oligodendrocyte progenitor (OP) cells. Activation of glutamate receptor channels by kainate, as well as stimulation of G-protein-coupled cholinergic receptors by carbachol and tyrosine kinase receptors by basic fibroblast growth factor (bFGF), rapidly leads to mitogen-activated protein kinase (MAPK) phosphorylation and ribosomal S6 kinase (RSK) activation. Kainate and carbachol activation of the MAPK pathway requires extracellular calcium influx and is accompanied by protein kinase C (PKC) induction, with no significant increase in GTP binding to Ras. Conversely, growth factor-stimulated MAPK phosphorylation is independent of extracellular calcium and is accompanied by Ras activation. Both basal and stimulated MAPK activity in OP cells are influenced by cytoplasmic calcium levels, as shown by their sensitivity to the calcium chelator bis(2-aminophenoxy)ethane-N,N,N',N'-tetra-acetic acid. The kinetics of CREB phosphorylation in response to the various agonists corresponds to that of MAPK activation. Moreover, CREB phosphorylation and MAPK activation are similarly affected by calcium ions. The MEK inhibitor PD 098059, which selectively prevents activation of the MAPK pathway, strongly reduces induction of CREB phosphorylation by kainate, carbachol, bFGF, and the phorbol ester TPA. We propose that in OPs the MAPK/RSK pathway mediates CREB phosphorylation in response to calcium influx, PKC activation, and growth factor stimulation.
The mitogen-activated protein kinases (MAPKs) ERK-1 and ERK-2 are activated by a wide variety of oncogenes and extracellular stimuli. The MAPKs participate in a signalling cascade downstream of growth factor/cytokine receptors, Ras, Raf, and MEK. However, MAPK activation is more complicated than a simple linear pathway, and the evidence presented here supports a model of multiple, temporally distinct pathways converging on MAPK which are differentially utilized by various stimuli and cell types. In addition to MEK-dependent MAPK activation, we provide evidence for MEK-independent regulation of the MAPKs. Our results suggest that phosphatidylinositol-3-kinases (PI(3)K) or conventional protein kinase C isoforms (cPKCs) partially contribute to MEK-dependent activation. Importantly, we also find that PI3K and cPKCs play a major role in the MEK-independent, prolonged MAPK activation by platelet-derived growth factor signalling. This finding is of interest as the maintained activation of MAPK has been correlated by others to the regulation of cell proliferation and differentiation.
Severely elevated levels of total homocysteine (approximately millimolar) in the blood typify the childhood disease homocystinuria, whereas modest levels (tens of micromolar) are commonly found in adults who are at increased risk for vascular disease and stroke. Activation of the coagulation system and adverse effects of homocysteine on the endothelium and vessel wall are believed to underlie disease pathogenesis. Here we show that homocysteine acts as an agonist at the glutamate binding site of the N-methyl-D-aspartate receptor, but also as a partial antagonist of the glycine coagonist site. With physiological levels of glycine, neurotoxic concentrations of homocysteine are on the order of millimolar. However, under pathological conditions in which glycine levels in the nervous system are elevated, such as stroke and head trauma, homocysteine's neurotoxic (agonist) attributes at 10-100 microM levels outweigh its neuroprotective (antagonist) activity. Under these conditions neuronal damage derives from excessive Ca2+ influx and reactive oxygen generation. Accordingly, homocysteine neurotoxicity through overstimulation of N-methyl-D-aspartate receptors may contribute to the pathogenesis of both homocystinuria and modest hyperhomocysteinemia.
In cell culture systems, the TCF Elk-1 represents a convergence point for extracellular signal-related kinase (ERK) and c-Jun N-terminal kinase/stress-activated protein kinase (JNK/SAPK) subclasses of mitogen-activated protein kinase (MAPK) cascades. Its phosphorylation strongly potentiates its ability to activate transcription of the c-fos promoter through a ternary complex assembled on the c-fos serum response element. In rat brain postmitotic neurons, Elk-1 is strongly expressed (V. Sgambato, P. Vanhoutte, C. Pagès, M. Rogard, R. A. Hipskind, M. J. Besson, and J. Caboche, J. Neurosci. 18:214-226, 1998). However, its physiological role in these postmitotic neurons remains to be established. To investigate biochemically the signaling pathways targeting Elk-1 and c-fos in mature neurons, we used a semi-in vivo system composed of brain slices stimulated with the excitatory neurotransmitter glutamate. Glutamate treatment leads to a robust, progressive activation of the ERK and JNK/SAPK MAPK cascades. This corresponds kinetically to a significant increase in Ser383-phosphorylated Elk-1 and the appearance of c-fos mRNA. Glutamate also causes increased levels of Ser133-phosphorylated cyclic AMP-responsive element-binding protein (CREB) but only transiently relative to Elk-1 and c-fos. ERK and Elk-1 phosphorylation are blocked by the MAPK kinase inhibitor PD98059, indicating the primary role of the ERK cascade in mediating glutamate signaling to Elk-1 in the rat striatum in vivo. Glutamate-mediated CREB phosphorylation is also inhibited by PD98059 treatment. Interestingly, KN62, which interferes with calcium-calmodulin kinase (CaM-K) activity, leads to a reduction of glutamate-induced ERK activation and of CREB phosphorylation. These data indicate that ERK functions as a common component in two signaling pathways (ERK/Elk-1 and ERK/?/CREB) converging on the c-fos promoter in postmitotic neuronal cells and that CaM-Ks act as positive regulators of these pathways.
Activation of the mitogen-activated protein kinase (MAPK) cascade recently was discovered to play an important role in synaptic plasticity in area CA1 of rat hippocampus. However, the upstream mechanisms regulating MAPK activity and the downstream effectors of MAPK in the hippocampus are uncharacterized. In the present studies we observed that hippocampal MAPK activation is regulated by both the PKA and PKC systems; moreover, we found that a wide variety of neuromodulatory neurotransmitter receptors (metabotropic glutamate receptors, muscarinic acetylcholine receptors, dopamine receptors, and beta-adrenergic receptors) couple to MAPK activation via these two cascades. In additional studies we observed that PKC is a powerful regulator of CREB phosphorylation in area CA1. MAPK plays a critical role in transcriptional regulation by PKC, because MAPK activation is a necessary component for increased CREB phosphorylation in response to the activation of this kinase. Surprisingly, we also observed that MAPK activation is necessary for PKA coupling to CREB phosphorylation in area CA1. Overall, these studies indicate an unexpected richness of diversity in the regulation of MAPK in the hippocampus and suggest the possibility of a broad role for the MAPK cascade in regulating gene expression in long-term forms of hippocampal synaptic plasticity.
Oxidative stress is implicated in the nerve cell death that occurs in a variety of neurological disorders, and the loss of protein kinase C (PKC) activity has been coupled to the severity of the damage. The functional relationship between stress, PKC, and cell death is, however, unknown. Using an immortalized hippocampal cell line that is particularly sensitive to oxidative stress, I show that activation of PKC by the phorbol ester tetradecanoylphorbol acetate (TPA) inhibits cell death via the stimulation of a complex protein phosphorylation pathway. TPA treatment leads to the rapid activation of extracellular signal-regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK), the inactivation of p38 mitogen-activated protein kinase (MAPK), and the downregulation of PKCdelta. Inhibition of either ERK or JNK activation blocks TPA-mediated protection, whereas p38 MAPK and PKCdelta inhibitors block stress-induced nerve cell death. Both p38 MAPK inactivation and JNK activation appear to be downstream of ERK because an agent that blocks ERK activation also blocks the modulation of these other MAP kinase family members by TPA treatment. Thus, the protection from oxidative stress afforded nerve cells by PKC activity requires the combined modulation of multiple enzyme pathways and suggests why the loss of PKC activity contributes to nerve cell death.
Basic fibroblast growth factor (FGF2) stimulates photoreceptor survival in vivo and in vitro, but the molecular signaling mechanism(s) involved are unknown. Immunohistochemical and immunoblotting analyses of pure photoreceptors,
inner retinal neurons, and Müller glial cells (MGC) in vitro revealed differential expression of the high affinity FGF receptors (FGFR1–4), as well as many cytoplasmic signaling intermediates
known to mediate the extracellular signal-regulated kinase (ERK1/2) pathway. FGF2-induced tyrosine phosphorylation in vitro exhibited distinct profiles for each culture type, and FGF2-induced ERK1/2 activation was observed for all three preparations.
Whereas U0126, a specific inhibitor of ERK kinase (MEK), completely abolished FGF2-induced ERK1/2 tyrosine phosphorylation
and survival in cultured photoreceptors, persistent ERK1/2 phosphorylation was observed in cultured inner retinal cells and
MGC. Furthermore U0126 treatment entirely blocked nerve growth factor-induced ERK1/2 activation in MGC, as well as FGF2-induced
ERK1/2 activation in cerebral glial cells. Taken together, these data indicate that FGF2-induced ERK1/2 activation is entirely
mediated by MEK within photoreceptors, which is responsible for FGF2-stimulated photoreceptor survival. In contrast, inner
retina/glia possess alternative, cell type, and growth factor-specific MEK-independent ERK1/2 activation pathways. Hence signaling
and biological effects elicited by FGF2 within retina are mediated by cell type-specific pathways.
Moderate hyperhomocysteinemia is associated with several diseases, including coronary artery disease, stroke, Alzheimer's disease, schizophrenia, and spina bifida. However, the mechanisms for their pathogenesis are unknown but could involve the interaction of homocysteine or its metabolites with molecular targets such as neurotransmitter receptors, channels, or transporters. We discovered that L-homocysteine sulfinic acid (L-HCSA), L-homocysteic acid, L-cysteine sulfinic acid, and L-cysteic acid are potent and effective agonists at several rat metabotropic glutamate receptors (mGluRs). These acidic homocysteine derivatives 1) stimulated phosphoinositide hydrolysis in the cells stably expressing the mGluR1, mGluR5, or mGluR8 (plus Galpha(qi9)) and 2) inhibited the forskolin-induced cAMP accumulation in the cells stably expressing mGluR2, mGluR4, or mGluR6, with different potencies and efficacies depending on receptor subtypes. Of the four compounds, L-HCSA is the most potent agonist at mGluR1, mGluR2, mGluR4, mGluR5, mGluR6, and mGluR8. The effects of the four agonists were selective for mGluRs because activity was not discovered when L-HCSA and several other homocysteine derivatives were screened against a large panel of cloned neurotransmitter receptors, channels, and transporters. These findings imply that mGluRs are candidate G-protein-coupled receptors for mediating the intracellular signaling events induced by acidic homocysteine derivatives. The relevance of these findings for the role of mGluRs in the pathogenesis of homocysteine-mediated phenomena is discussed.
Cystathionine beta-synthase (CBS) deficiency causes severe hyperhomocysteinemia and other signs of homocystinuria syndrome, in particular a premature atherosclerosis with multiple thrombosis. However, the molecular mechanisms by which homocysteine could interfere with normal cell function are poorly understood in a whole organ like the liver, which is central to the catabolism of homocysteine. We used a combination of differential display and cDNA arrays to analyze differential gene expression in association with elevated hepatic homocysteine levels in CBS-deficient mice, a murine model of hyperhomocysteinemia. Expression of several genes was found to be reproducibly abnormal in the livers of heterozygous and homozygous CBS-deficient mice. We report altered expression of genes encoding ribosomal protein S3a and methylthioadenosine phosphorylase, suggesting such cellular growth and proliferation perturbations may occur in homozygous CBS-deficient mice liver. Many up- or down-regulated genes encoded cytochromes P450, evidence of perturbations of the redox potential in heterozygous and homozygous CBS-deficient mice liver. The expression of various genes involved in severe oxidative processes was also abnormal in homozygous CBS-deficient mice liver. Among them, the expression of heme oxygenase 1 gene was increased, concomitant with overexpression of heme oxygenase 1 at the protein level. Commensurate with the difference in hepatic mRNA paraoxonase 1 abundance, the mean hepatic activity of paraoxonase 1, an enzyme that protects low density lipoprotein from oxidation, was 3-fold lower in homozygous CBS-deficient mice. Heterozygous CBS-deficient mice, when fed a hyperhomocysteinemic diet, have also reduced PON1 activity, which demonstrates the effect of hyperhomocysteinemia in the paraoxonase 1 activity.
Homocystinuria with elevated plasma homocysteine and methionine levels is the result of deficient activity of cystathionine synthetase, the enzyme catalyzing conversion of homocysteine to cystathionine. It is inherited as an autosomal recessive trait with a worldwide distribution. The major clinical manifestations result from the elevated plasma homocysteine level. The excitotoxic effect of homocysteic acid accounts for mental retardation and seizures. Interference with collagen cross-linking by sulfhydryl groups of homocysteine causes ectopia lentis and skeletal deformities. Sulfation factor-like effects contribute to disruption of vascular endothelium, which is followed by platelet thrombosis and widespread arterial and venous occlusions. Low methionine homocystinuria, with deficient remethylation of homocysteine, results from deranged vitamin B12 metabolism and from deficient 5,10-methylene-tetrahydrofolate reductase. Administration of azaribine produces homocystinuria by a mechanism not yet elucidated.
Biological thiols can regulate cell signal transduction. The effects of two biothiols, (Hcy), a risk factor for cardiovascular disease, and alpha-lipoic acid (alpha LA), a therapeutic antioxidant, on p44/42 mitogen-activated protein kinases (MAPK) phosphorylation were examined in NIH/3T3 fibroblasts. Cells grown in serum-containing media had constitutive levels of MAPK phosphorylation as determined by Western blot analysis using the phospho-specific MAPK antibody. Treatment of cells with 20 mu M Hcy for 0-60 min resulted in a transient enhancement of MAPK phosphorylation. In contrast, 20 mu M alpha LA inhibited serum-mediated phosphorylation of MAPK. The differential effects of these two thiols are not due to their redox states as oxidized Hcy ( Hcy thiolactone) enhanced MAPK phosphorylation. The effect of alpha LA appears to be serum-dependent because Hcy or alpha LA treatment of serum-deprived cells activated MAPK phosphorylation. Thus, alpha LA and Hcy can either induce common signal transduction pathways or differentially modulate MAPK phosphorylation, depending on the state of the cell. This relationship may be important to understand how some biothiols are associated with pathogenic events while others offer potential as therapeutic agents. Antiox. Redox Signal. 1, 123-128.
Stress-activated protein kinase (SAPK) and extracellular signal-regulated kinase (ERK), both members of the mitogen-activated protein kinase (MAPK) family, may in some circumstances serve opposing functions with respect to cell survival. However, SAPK and ERK can also be coordinately activated in neurons in response to glutamate stimulation of NMDA receptors. To explore the mechanisms of these MAPK activations, we compared the ionic mechanisms mediating SAPK and ERK activations by glutamate. In primary cultures of striatal neurons, glutamatergic activation of ERK and one of its transcription factor targets, CREB, showed a calcium dependence typical of NMDA receptor-mediated responses. In contrast, extracellular calcium was not required for glutamatergic, NMDA receptor-mediated activation of SAPK and phosphorylation of its substrate, c-Jun. Increasing extracellular calcium enhanced ERK activation but reversed SAPK activation, further distinguishing the calcium dependencies of these two NMDA receptor-mediated effects. Finally, reducing extracellular sodium prevented the glutamatergic activation of SAPK but only partially blocked that of ERK. These contrasting ionic dependencies suggest a mechanism by which NMDA receptor activation may, under distinct conditions, differentially regulate neuronal MAPKs and their divergent functions.
BACKGROUND: Alteration in mesangial cell function is central to the progression of glomerular disease in numerous models of chronic renal failure (CRF). Animal models of chronic glomerular disease are characterized by mesangial cell proliferation and elaboration of extracellular matrix protein (ECM), resulting in glomerulosclerosis. Elevated plasma levels of homocysteine (Hcy) are seen in both animal models and humans with CRF, and have been proposed to contribute to the high prevalence of vascular disease in this group. Some of the pathogenetic effects of Hcy are thought to be mediated via the induction of endoplasmic reticulum stress. Thus, Hcy effects on mesangial cells could contribute to the progression of CRF. Previous work has shown Hcy- mediated induction of Erk mitogen-activated protein kinase (MAPK) in vascular smooth muscle cells (VSMCs). Erk induces increases in activator protein-1 (AP-1) transcription factor activity which may augment mesangial cell proliferation and ECM protein production. Consequently, we studied the effect of Hcy on mesangial cell Erk signaling. METHODS: Mesangial cells were exposed to Hcy after 24 hours of serum starvation and Erk activity assessed. Nuclear translocation of phospho-Erk was visualized by confocal microscopy. AP-1 nuclear protein binding was measured in response to Hcy by mobility shift assay. Hcy-induced mesangial cell calcium flux was measured in Fura-2 loaded cells. Mesangial cell DNA synthesis in response to Hcy was assessed by [3H]-thymidine incorporation and proliferation by Western blotting for proliferating cell nuclear antigen (PCNA). Expression of endoplasmic reticulum stress response genes were determined by Northern and Western analysis. RESULTS: Hcy led to an increase in Erk activity that was maximal at 50 micromol/L and 20 minutes of treatment. Subsequent experiments used this concentration and time point. Erk activity in response to Hcy was insensitive to n-acetylcysteine and catalase, indicating oxidative stress did not play a role. However, Hcy50 micromol/L induced a brief increase in intracellular mesangial cell calcium within 5 minutes, and the calcium ionophores A23187 and ionomycin increased Erk activity while chelation of intracellular calcium with BAPTA-AM abrogated the Erk response to Hcy. Confocal microscopy of activated Erk nuclear translocation mirrored these results as did mesangial cell nuclear protein binding to AP-1 consensus sequences. Hcy- induced increases in thymidine incorporation and PCNA expression at 24 hours were Erk dependent. The expression of endoplasmic reticulum stress response genes was significantly elevated by Hcy in an Erk-dependent manner. CONCLUSION: Hcy increases Erk activity in mesangial cells via a calcium-dependent mechanism, resulting in increased AP-1 nuclear protein binding, cell DNA synthesis and proliferation and induction of endoplasmic reticulum stress. These observations suggest potential mechanisms by which Hcy may contribute to progressive glomerular injury.
Homocystinuria with elevated plasma homocysteine and methionine levels is the result of deficient activity of cystathionine synthetase, the enzyme catalyzing conversion of homocysteine to cystathionine. It is inherited as an autosomal recessive trait with a worldwide distribution. The major clinical manifestations result from the elevated plasma homocysteine level. The excitotoxic effect of homocysteic acid accounts for mental retardation and seizures. Interference with collagen cross-linking by sulfhydryl groups of homocysteine causes ectopia lentis and skeletal deformities. Sulfation factor-like effects contribute to disruption of vascular endothelium, which is followed by platelet thrombosis and widespread arterial and venous occlusions. Low methionine homocystinuria, with deficient remethylation of homocysteine, results from deranged vitamin B(12) metabolism and from deficient 5,10-methylene-tetrahydrofolate reductase. Administration of azaribine produces homocystinuria by mechanism not yet elucidated.
The transport kinetics of the excitatory sulphur-containing amino acid (SAA) transmitter candidates, L-cysteine sulphinate (L-CSA), L-cysteate (L-CA), L-homocysteine sulphinate (L-HCSA), and L-homocysteate (L-HCA), together with their plasma membrane carrier specificity, was studied in cerebrocortical synaptosome fractions by a sensitive high performance liquid chromatographic assay. A high affinity uptake system could be demonstrated for L-CSA (Km = 57 +/- 6 microM; Vmax = 1.2 +/- 0.1 nmol/min/mg protein) and L-CA (Km = 23 +/- 3 microM; Vmax = 3.6 +/- 0.1 nmol/min/mg protein), whereas L-HCSA (Km = 502 +/- 152 microM; Vmax = 6.1 +/- 1.3 nmol/min/mg protein) and L-HCA (Km = 1550 +/- 169 microM; Vmax = 10.3 +/- 1.1 nmol/min/mg protein) exhibited much lower affinity as transport substrates. In all cases, only a single, saturable Na(+)-dependent component of uptake could be identified, co-existing with a non-saturable, Na(+)-independent influx component. Plasma membrane carrier specificity of the SAAs was established following comparison with other high-affinity neurotransmitter systems. High-affinity L-CSA and L-CA transport and low-affinity L-HCSA and L-HCA transport demonstrate strong positive correlations in inhibition profiles when compared against each other or individually against the high-affinity transport of L-[3H]glutamate, L-[3H]aspartate, or D-[3H]aspartate. Moreover, the transport systems for the excitatory SAAs exhibited a negative correlation when compared in inhibition profiles with the high affinity transport of both [3H] gamma-aminobutyric acid (GABA) and [3H]taurine.(ABSTRACT TRUNCATED AT 250 WORDS)
Sulphur-containing excitatory amino acid transmitter candidates (500 microM) stimulated the Ca(2+)-independent efflux of exogenously-supplied D-[3H]aspartate from primary cultures of cerebellar granule cells superfused continuously with HEPES-buffered saline containing CoCl2 (1 mM) in place of CaCl2. The stimulated release of D-[3H]aspartate was markedly attenuated by 200 microM 6,7-dinitroquinoxalinedione, a concentration at which the antagonist inhibits both non-N-methyl-D-aspartate and N-methyl-D-aspartate ionotropic excitatory amino acid receptors. The Ca(2+)-independent component of evoked release was also markedly attenuated and, in some cases, abolished by removing NaCl from the superfusion medium. Furthermore, when 700 microM dihydrokainate (demonstrated herein as a mixed/non-competitive inhibitor of the high-affinity dicarboxylic amino acid transporter in cultured granule cells) was included in the superfusion medium, stimulated efflux of D-[3H]aspartate was reduced by between 15-78% of the control response; the extent of inhibition varying with the agonist employed. In constrast, agents which act as competitive inhibitors of the plasma membrane carrier in granule cells, e.g. beta-methylene-D,L-aspartate, potentiated the release of D-[3H]aspartate in a synergistic manner. Taken together, these findings are consistent with a mechanism for the Ca(2+)-independent release of D-[3H]aspartate that is mediated predominantly by activation of excitatory amino acid receptors resulting in a reversal of the high-affinity dicarboxylic amino acid transport system. Although the physiological relevance of such non-vesicular release from the cytosol remains obscure and is still a matter of some debate, this mode of release may be of pathological significance.
A detailed kinetic study of the inhibitory effects ofl- andd-enantiomers of cysteate, cysteine sulphinate, homocysteine sulphinate, homocysteate, and S-sulpho-cysteine on the neuronal, astroglial and synaptosomal high-affinity glutamate transport system was undertaken.d-[3H] Aspartate was used as the transport substrate. Kinetic characterisation of uptake in the absence of sulphur compounds confirmed the high-affinity nature of the transport systems, the Michaelis constant (K
m) ford-aspartate uptake being 6 μM, 21 μM and 84 μM, respectively, in rat brain cortical synaptosomes and primary cultures of mouse cerebellar granule cells and cortical astrocytes. In those cases where significant effects could be demonstrated, the nature of the inhibition was competitive irrespective of the neuronal versus glial systems. The rank order of inhibition was essentially similar in synaptosomes, neurons and astrocytes. Potent inhibition (K
i∼K
m) of transport in each system was exhibited byl-cysteate, andl- andd-cysteine sulphinate whereas substantially weaker inhibitory effects (K
i>10–1000 times the appropriateK
m value) were exhibited by the remaining sulphur amino acids. In general, inhibition: (i) was markedly stereospecific in favor of thel-enantiomers (except for cysteine sulphinate) and (ii) was found to decrease with increasing chain length. Computer-assisted molecular modelling studies, in which volume contour maps of the sulphur compounds were superimposed on those ofd-aspartate andl-glutamate, demonstrated an order of inhibitory potency which was, qualitatively, in agreement with that obtained quantitatively by in vitro kinetic studies.
An international questionnaire survey has been conducted to define better the natural history of homocystinuria due to cystathionine beta-synthase deficiency and permit evaluation of treatment. Data were compiled for 629 patients. Among patients not discovered by newborn screening, B6-responsive individuals on the average have significantly better mental capabilities (mean IQ, 79) than do B6-nonresponsive individuals (mean IQ, 57). Time-to-event curves are presented for the other major clinical abnormalities produced by this disease. Each occurred at significantly lower rates in untreated B6-responsive than in untreated B6-nonresponsive patients, as shown by the following examples: (1) dislocation of optic lenses (at age 10, chances of dislocation: 55% and 82%, respectively); (2) initial clinically detected thromboembolic events (at age 15, chances of having had such an event: 12% and 27%, respectively); (3) radiologic detection of spinal osteoporosis (at age 15, chances of such osteoporosis having been detected: 36% and 64%, respectively); and (4) mortality (at age 30, chances of not surviving: 4% and 23%, respectively). Methionine restriction initiated neonatally prevented mental retardation, retarded the rate of lens dislocation, and may have reduced the incidence of seizures. Pyridoxine treatment of late-detected B6-responsive patients retarded the rate of occurrence of initial thromboembolic events. Following 586 surgical procedures, 25 postoperative thromboembolic complications occurred, six of which were fatal. Reproductive histories were reported predominantly for B6-responsive patients. Living offspring of either men or women patients had few abnormalities. The evidence is inconclusive whether untreated maternal cystathionine beta-synthase deficiency leads to excessive fetal loss. Only 13% of patients detected in screening programs of newborns and classified as to B6-responsiveness were B6-responsive, compared to 47% among late-detected patients. Current screening programs that identify neonatal hypermethioninemia may be preferentially failing to detect B6-responsive patients.
Severe or even mild hyperhomocysteinaemia can cause a wide range of neurological problems. In recent years its vascular complications, including cerebral stroke, in children and young adults have gained special interest, because hyperhomocysteinaemia is treatable and recurrence of vascular incidents may be preventable. Current knowledge about biochemical mechanisms leading to hyperhomocysteinaemia, the pathogenesis of vascular pathology and neurological disfunction, and the various patterns of cerebral damage are reviewed. The significance of MRI in diagnosis, follow-up and research on hyperhomocysteinaemia is discussed.
Repetitive activation of hippocampal mossy fibers evokes a long-term potentiation (LTP) of synaptic responses in pyramidal
cells in the CA3 region that is independent of N-methyl-D-aspartate receptor activation. Previous results suggest that the
site for both the induction and expression of this form of LTP is presynaptic. Experimental elevation of cyclic adenosine
3',5'-monophosphate (cAMP) both mimics and interferes with tetanus-induced mossy fiber LTP, and blockers of the cAMP cascade
block mossy fiber LTP. It is proposed that calcium entry into the presynaptic terminal may activate Ca(2+)-calmodulin-sensitive
adenylyl cyclase I which, through protein kinase A, causes a persistent enhancement of evoked glutamate release.
Ca2+ influx through N-methyl-D-aspartate- (NMDA-) type glutamate receptors plays a critical role in synaptic plasticity in the brain. One of the proteins activated by the increase in Ca2+ is CaM kinase II (CaMKII). Here, we report a novel synaptic Ras-GTPase activating protein (p135 SynGAP) that is a major component of the postsynaptic density, a complex of proteins associated with synaptic NMDA receptors. p135 SynGAP is almost exclusively localized at synapses in hippocampal neurons where it binds to and closely colocalizes with the scaffold protein PSD-95 and colocalizes with NMDA receptors. The Ras-GTPase activating activity of p135 SynGAP is inhibited by phosphorylation by CaMKII located in the PSD protein complex. Inhibition of p135 SynGAP by CaMKII will stop inactivation of GTP-bound Ras and thus could result in activation of the mitogen-activated protein (MAP) kinase pathway in hippocampal neurons upon activation of NMDA receptors.
Homocysteine at abnormally high levels is an independent risk factor for atherosclerosis and may be a key factor in atherogenesis. Since homocysteine (Hcys) has been shown to promote cell proliferation and induction of the gene transcription factor c-fos in vascular smooth muscle cells (VSMCs), effects which can be mediated by MAP kinase, we hypothesized that homocysteine activates a MAP kinase-dependent signal transduction pathway. In this study, we find that homocysteine transiently activates MAP kinase (ERK2 isoform) in cultured VSMCs from chick embryos. Homocysteine activation of ERK2 is dose-dependent with an EC50 of approximately 500 nM and blocked by the MAP/Erk kinase (MEK) inhibitor PD98059. VSMC embryonic lineage is another determinant of homocysteine sensitivity. These findings demonstrate that homocysteine activates the MAP kinase signal transduction pathway and thus support the hypothesis that homocysteine may promote atherosclerosis by stimulation of growth promoting signal transduction pathways.
Mitogen-activated protein kinases (MAPKs) are activated by various stimuli, such as growth factors, cytokines, or stress, and are considered to be important mediators in intracellular signal transduction networks. The dual-specificity kinases, MAPK kinases (MKKs), which phosphorylate the TXY motif in the catalytic domain of MAPKs, can cause the activation of MAPKs. Recently, a family of dual-specificity phosphatases has been identified, members of which are able to dephosphorylate and inactivate MAPKs. The studies cited in this review have revealed that these MAPK phosphatases might play an important role in various cellular functions by downregulating the MAPK cascade.
Homocysteine (Hcy) exerts either promoting or suppressive effects on mitogenesis in a cell type-specific manner. Hcy elicits proliferation of vascular smooth muscle cells, but is rather inhibitory to growth of endothelial cells and NIH/3T3 cells. In NIH/3T3 cells, we found that physiologically relevant concentrations (20-100 microM) of Hcy inhibit the activity of activating protein-1 (AP-1) transcription factor, although it is capable of eliciting immediate-early signaling events. Hcy induced p44/42 mitogen-activated protein kinase (MAPK) phosphorylation in control cells, but not in dominant negative p21ras transfected cells, indicating induction of the Ras-MAPK pathway. Hcy also induced the activity of serum response factor and expression of c-fos and c-jun genes. Despite the activation of these upstream events, Hcy potently inhibited AP-1 activity. Oxidized forms of Hcy (Hcy thiolactone, homocystine) were less effective in affecting AP-1. Hcy-mediated inhibition of AP-1 activity was not observed in A7r5 vascular smooth muscle cells. These results demonstrate that Hcy exerts cell type- and redox-specific inhibition of AP-1 dependent biological events.
In vivo activation of group I metabotropic glutamate receptors (mGluRs) upregulates phosphorylation of cyclic AMP response element-binding protein (CREB), Elk-1 and extracellular signal-regulated kinases (ERK) in striatal neurons. To evaluate putative roles of Ca2+/calmodulin-dependent protein kinase II (CaMKII) in CREB, Elk-1 and ERK phosphorylation, the CaMKII inhibitor, KN62, was infused simultaneously with the group I mGluR agonist, 3,5-dihydroxyphenylglycine (DHPG), into the rat dorsal striatum. The results showed that DHPG (125, 250, and 500 nmol) increased phosphorylated (p) CaMKII immunoreactivity (IR) in a dose-dependent manner. KN62 (50 nmol) significantly attenuated 500 nmol DHPG-induced pERK, pElk-1 and pCREB IR in the ipsilateral dorsal striatum. These data indicate that pCaMKII is a possible upstream effector that is responsible for the regulation of CREB, Elk-1 and ERK phosphoproteins in response to group I mGluR stimulation in striatal neurons.
Metabotropic glutamate receptors (mGluRs) are implicated in the regulation of diverse neuronal plasticity and neuropathological processes in the central nervous system. Activation of mGluRs couples glutamatergic signals to second messengers in a subtype-specific manner: activation of group I mGluRs upregulates Ca2+ cascades, while group II/III downregulates the adenylate cyclase and cAMP cascades. Dominant presynaptic inhibitory actions of group II/III mGluRs on the glutamate release, extensive cross-talk between kinases by various second messengers downstream to the group I mGluRs, and desensitization of mGluRs in response to prolonged stimulation of glutamate input have been documented in the regulation of glutamatergic transmission. In addition to the spatiotemporal processes, interactions with ionotropic glutamate receptors, and protein phosphatase activity against kinase actions further regulate glutamatergic signals. These overall activities in medium spiny neurons contribute to modifying striatal outflow in striatopallidal and striatonigral neurons. Thus, characterization of the roles of mGluRs in the regulation of intracellular effectors is crucial for the understanding of diverse neuronal plasticity implicated with the receptors including long-term potentiation and long-term depression, neurotoxicity, actions of abused drugs, and neurodegenerative diseases. In this review we attempted to provide a broad spectrum on how mGluRs regulate the phosphorylation of cAMP response element-binding protein and Elk-1, well known inducible transcription factors by extracellular stimuli, by emphasizing major kinase interactions in medium spiny neurons.
The mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) pathway is an evolutionarily conserved signaling cascade involved in a plethora of physiological responses, including cell proliferation, survival, differentiation, and, in neuronal cells, synaptic plasticity. Increasing evidence now implicates this pathway in cognitive functions, such as learning and memory formation, and also in behavioral responses to addictive drugs. Although multiple intracellular substrates can be activated by ERKs, nuclear targeting of transcription factors, and thereby control of gene expression, seems to be a major event in ERK-induced neuronal adaptation. By controlling a prime burst of gene expression, ERK signaling could be critically involved in molecular adaptations that are necessary for long-term behavioral changes. Reviewed here are data providing evidence for a role of ERKs in long-term behavioral alterations, and the authors discuss molecular mechanisms that could underlie this role.
The authors examined the association of total plasma homocysteine (Hcy) levels with measures of atrophy and white matter disease on MRI scans in 36 healthy elderly individuals. Hcy had a significant positive relationship with lateral ventricle-brain ratios in the anterior (r = 0.49) and middle (r = 0.43) ventricular regions as measures of central atrophy, but not with cortical atrophy or white matter hyperintensities. In a logistic regression analysis, elevated Hcy was a significant determinant of increased anterior ventricle-brain ratio (> or =0.34) after controlling for age, folate, B12, creatinine, and white matter disease (OR = 2.3; CI, 1.03-5.09).
Phorbol esters are hypothesised to produce a protein kinase C (PKC)-dependent increase in the probability of transmitter release via two mechanisms: facilitation of vesicle fusion or increases in synaptic vesicle number and replenishment. We used a combination of electrophysiology and computer simulation to distinguish these possibilities. We constructed a stochastic model of the presynaptic contacts between a pair of hippocampal pyramidal cells that used biologically realistic processes and was constrained by electrophysiological data. The model reproduced faithfully several forms of short-term synaptic plasticity, including short-term synaptic depression (STD), and allowed us to manipulate several experimentally inaccessible processes. Simulation of an increase in the size of the readily releasable vesicle pool and the time of vesicle replenishment decreased STD, whereas simulation of a facilitation of vesicle fusion downstream of Ca(2+) influx enhanced STD. Because activation of protein kinase C with phorbol ester enhanced STD of EPSCs in rat hippocampal slice cultures, we conclude that an increase in the sensitivity of the release process for Ca(2+) underlies the potentiation of neurotransmitter release by PKC.
Homocysteine (HC) is a neurotoxic amino acid that accumulates in several neurological disorders including Alzheimer's disease (AD). We examined the consequences of treatment of cultured murine cortical neurons with HC. Homocysteine-induced increases in cytosolic calcium, reactive oxygen species, phospho-tau immunoreactivity and externalized phosphatidyl serine (indicative of apoptosis). Homocysteine-induced calcium influx through NMDA channel activation, which stimulated glutamate excitotoxicity, as evidenced by treatment with antagonists of the NMDA channel and metabotropic glutamate receptors, respectively. The NMDA channel antagonist MK-801 reduced tau phosphorylation but not apoptosis after HC treatment, suggesting that HC-mediated apoptosis was not due to calcium influx. Apoptosis after HC treatment was reduced by co-treatment with 3-aminobenazmidine (3ab), an inhibitor of poly-ADP-ribosome polymerase (PARP), consistent with previous reports that ATP depletion by PARP-mediated repair of DNA strand breakage mediated HC-induced apoptosis. Treatment with 3ab did not reduce tau phosphorylation, however, therefore hyperphosphorylation of tau may not contribute to HC-induced apoptosis under these conditions. Inhibition of mitogen-activated protein kinase by co-treatment with the kinase inhibitor PD98059 inhibited tau phosphorylation but not apoptosis after HC treatment. HC accumulation reduces cellular levels of S-adenosyl methionine (SAM); co-treatment with SAM reduced apoptosis, suggesting that inhibition of critical methylation reactions may mediate HC-induced apoptosis. These findings indicate that HC compromises neuronal homeostasis by multiple, divergent routes.
This in vivo microdialysis study compared the effects of NMDA and D,L-homocysteine (Hcy) administered via dialysis medium on 45Ca efflux from prelabeled rabbit hippocampus. Application of these agonists evoked dose-dependent, and sensitive to MK-801, opposite effects: NMDA decreased the 45Ca radioactivity in the dialysate, whereas Hcy induced the release of 45Ca. The latter effect was potentiated by glycine, inhibited by the antagonist of group I metabotropic glutamate receptors (mGluR) LY367385, and mimicked by t-ADA, an agonist of these receptors. Electron microscopic examination of pyramidal neurones in the CA1 sector of the hippocampus in the vicinity of the microdialysis probe after NMDA application demonstrated swelling of mitochondria, which was prevented by cyclosporin A. This study shows, for the first time, Hcy-induced activation of both group I mGluR and NMDA receptors, which may play a role in acute Hcy neurotoxicity. We present new applications of brain microdialysis in studies on excitotoxicity and neuroprotection.
Hyperhomocysteinemia is a risk factor in neurodegeneration. It has been suggested that apart from disturbances in methylation processes, the mechanisms of this effect may include excitotoxicity mediated by the N-methyl-D-aspartate (NMDA) receptors. In this study we demonstrate that apart from NMDA receptors, also group I metabotropic glutamate receptors participate in acute homocysteine (Hcy)-induced neurotoxicity in cultured rat cerebellar granule neurones. Primary neuronal cultures were incubated for 30 min in the Mg(2+)-free ionic medium containing homocysteine and other ligands, and neurodegenerative changes were assessed 24h later using propidium iodide staining. D,L-Homocysteine given alone appeared to be a weak neurotoxin, with EC(50) of 17.4mM, whereas EC(50) for L-glutamate was 0.17 mM. Addition of 50 microM glycine enhanced homocysteine neurotoxicity, and only that portion of neurotoxicity was abolished by 0.5 microM MK-801, an uncompetitive NMDA receptor antagonist. The net stimulation of 45Ca uptake by granule cells incubated in the presence of 25 mM D,L-homocysteine with 50 microM glycine was only 3% of the net uptake evoked by 1mM glutamate. Application of an antagonist of group I metabotropic glutamate receptors (mGluRs) LY367385 at 25 and 250 microM concentrations, induced a dose-dependent partial neuroprotection, whereas given together with MK-801 completely prevented neurotoxicity. In the absence of glycine, LY367385 and MK-801 given alone failed to induce neuroprotection, while applied together completely prevented homocysteine neurotoxicity. Agonist of group I mGluRs, 10 trans-azetidine-2,3-dicarboxylic acid (t-ADA) induced significant neurotoxicity. This study shows for the first time that acute homocysteine-induced neurotoxicity is mediated both by group I mGluRs and NMDA receptors, and is not accompanied by massive influx of extracellular Ca(2+) to neurones.
Clinical studies suggest a relationship between folate deficiency and neurological and disorders including Alzheimer's disease (AD). To investigate mechanisms underlying this association, we examined the consequences of folate deprivation on neuronal cultures. Culturing embryonic cortical neurons and differentiated SH-SY-5Y human neuroblastoma cells in folate-free medium induced neurodegenerative changes characteristic of those observed in AD, including increased cytosolic calcium, reactive oxygen species (ROS), phospho-tau and apoptosis. In accord with clinical studies, generation of the neurotoxic amino acid homocysteine (HC) was likely to contribute to these phenomena, since (1) a significant increase in HC was detected following folate deprivation, (2) addition of the inhibitor of HC formation, 3-deazaadenosine, both prevented HC formation and eliminated the increase in ROS that normally accompanied folate deprivation, (3) direct addition of HC in the presence of folate induced the neurotoxic effects that accompanied folate deprivation, and (4) an antagonist of NMDA channels that blocks HC-induced calcium influx also blocked calcium influx following folate deprivation. Folate deprivation decreased the reduced form of glutathione, indicating a depletion of oxidative buffering capacity. This line of reasoning was supported by an increase in glutathione and reduction in ROS following supplementation of folate-deprived cultures with the cell-permeant glutathione precursor, N-acetyl-L-cysteine, or vitamin E. Folate deprivation potentiated ROS and apoptosis induced by amyloid-beta, while folate supplementation at higher concentrations prevented generation of ROS by amyloid-beta, suggesting that folate levels modulate the extent of amyloid-beta neurotoxicity. These findings underscore the importance of folate metabolism in neuronal homeostasis and suggest that folate deficiency may augment AD neuropathology by increasing ROS and excitotoxicity via HC generation.
Glial cells synthesise neuroactive substances and release them upon neurotransmitter receptor activation. Homocysteic acid (HCA), an endogenous agonist for glutamatergic N-methyl-D-aspartate (NMDA) receptors, is predominantly localised in glial cells. We have previously demonstrated the release of HCA from mouse astrocytes in culture following activation of beta-adrenergic receptors. Moreover, a release of HCA has also been observed in vivo upon physiological stimulation of sensory afferents in the thalamus. Here we report the glutamate-induced release of HCA from astrocytes. The effect of glutamate was mediated by the activation of ionotropic (NMDA and non-NMDA) as well as by metabotropic receptors. In addition, the release of HCA was Ca(2+)- and Na(+)-dependent, and its mechanism involved the activation of the Na+/Ca(2+)-exchanger. Furthermore, we provide evidence for the presence of functional NMDA receptors on astrocytes, which are coupled to an intracellular Ca2+ increase via stimulation of the Na+/Ca(2+)-exchanger. Our data thus favour a participation of glial cells in excitatory neurotransmission and corroborate the role of HCA as a "gliotransmitter."
Deficiency in cystathionine beta synthase (CBS) leads to high plasma homocysteine concentrations and causes hyperhomocysteinemia, a common risk factor for vascular disease, stroke and possibly neurodegenerative diseases. Various neuronal diseases have been associated with hyperhomocysteinemia, but the molecular mechanisms of homocysteine toxicity are unknown. We investigated the pathways involved in the pathological process, by analyzing differential gene expression in neuronal tissues. We used a combination of differential display and cDNA arrays to identify genes differentially expressed during hyperhomocysteinemia in brain of CBS-deficient mice. In this murine model of hyperhomocysteinemia, both plasma and brain homocysteine concentrations were high. Several genes were found to be differentially expressed in the brains of CBS-deficient mice, and the identities of some of these genes suggested that the SAPK/JNK pathway was altered in the brains of CBS-deficient mice. We therefore investigated the activation of proteins involved in the SAPK/JNK cascade. JNK and c-Jun were activated in the hippocampal neurones of CBS-deficient mice, suggesting that the SAPK/JNK pathway may play an important role in the development of neuronal defects associated with hyperhomocysteinemia.
Alteration in mesangial cell function is central to the progression of glomerular disease in numerous models of chronic renal failure (CRF). Animal models of chronic glomerular disease are characterized by mesangial cell proliferation and elaboration of extracellular matrix protein (ECM), resulting in glomerulosclerosis. Elevated plasma levels of homocysteine (Hcy) are seen in both animal models and humans with CRF, and have been proposed to contribute to the high prevalence of vascular disease in this group. Some of the pathogenetic effects of Hcy are thought to be mediated via the induction of endoplasmic reticulum stress. Thus, Hcy effects on mesangial cells could contribute to the progression of CRF. Previous work has shown Hcy- mediated induction of Erk mitogen-activated protein kinase (MAPK) in vascular smooth muscle cells (VSMCs). Erk induces increases in activator protein-1 (AP-1) transcription factor activity which may augment mesangial cell proliferation and ECM protein production. Consequently, we studied the effect of Hcy on mesangial cell Erk signaling.
Mesangial cells were exposed to Hcy after 24 hours of serum starvation and Erk activity assessed. Nuclear translocation of phospho-Erk was visualized by confocal microscopy. AP-1 nuclear protein binding was measured in response to Hcy by mobility shift assay. Hcy-induced mesangial cell calcium flux was measured in Fura-2 loaded cells. Mesangial cell DNA synthesis in response to Hcy was assessed by [3H]-thymidine incorporation and proliferation by Western blotting for proliferating cell nuclear antigen (PCNA). Expression of endoplasmic reticulum stress response genes were determined by Northern and Western analysis.
Hcy led to an increase in Erk activity that was maximal at 50 micromol/L and 20 minutes of treatment. Subsequent experiments used this concentration and time point. Erk activity in response to Hcy was insensitive to n-acetylcysteine and catalase, indicating oxidative stress did not play a role. However, Hcy50 micromol/L induced a brief increase in intracellular mesangial cell calcium within 5 minutes, and the calcium ionophores A23187 and ionomycin increased Erk activity while chelation of intracellular calcium with BAPTA-AM abrogated the Erk response to Hcy. Confocal microscopy of activated Erk nuclear translocation mirrored these results as did mesangial cell nuclear protein binding to AP-1 consensus sequences. Hcy- induced increases in thymidine incorporation and PCNA expression at 24 hours were Erk dependent. The expression of endoplasmic reticulum stress response genes was significantly elevated by Hcy in an Erk-dependent manner.
Hcy increases Erk activity in mesangial cells via a calcium-dependent mechanism, resulting in increased AP-1 nuclear protein binding, cell DNA synthesis and proliferation and induction of endoplasmic reticulum stress. These observations suggest potential mechanisms by which Hcy may contribute to progressive glomerular injury.
Increased serum level of homocysteine, a sulfur-containing amino acid, is considered a risk factor in vascular disorders and in dementias. The effect of homocysteine and metabolically related compounds on brain production of kynurenic acid (KYNA), an endogenous antagonist of glutamate ionotropic receptors, was studied. In rat cortical slices, DL-homocysteine enhanced (0.1-0.5 mM) or inhibited (concentration inducing 50% inhibition [IC50]=6.4 [5.5-7.5] mM) KYNA production. In vivo peripheral application of DL-homocysteine (1.3 mmol/kg intraperitoneally) increased KYNA content (pmol/g tissue) from 8.47 +/- 1.57 to 13.04 +/- 2.86 (P <0.01; 15 min) and 11.4 +/- 1.72 (P <0.01; 60 min) in cortex, and from 4.11 +/- 1.54 to 10.02 +/- 3.08 (P <0.01; 15 min) in rat hippocampus. High concentrations of DL-homocysteine (20 mM) applied via microdialysis probe decreased KYNA levels in rabbit hippocampus; this effect was antagonized partially by an antagonist of group I metabotropic glutamate receptors, LY367385. In vitro, S-adenosylhomocysteine acted similar to but more potently than DL-homocysteine, augmenting KYNA production at 0.03-0.08 mM and reducing it at > or =0.5 mM. The stimulatory effect of S-adenosylhomocysteine was abolished in the presence of the L-kynurenine uptake inhibitors L-leucine and L-phenyloalanine. Neither the N-methyl-D-aspartate (NMDA) antagonist CGS 19755 nor L-glycine influenced DL-homocysteine- and S-adenosylhomocysteine-induced changes of KYNA synthesis in vitro. DL-Homocysteine inhibited the activity of both KYNA biosynthetic enzymes, kynurenine aminotransferases (KATs) I and II, whereas S-adenosylhomocysteine reduced only the activity of KAT II. L-Methionine and L-cysteine, thiol-containing compounds metabolically related to homocysteine, acted only as weak inhibitors, reducing KYNA production in vitro and inhibiting the activity of KAT II (L-cysteine) or KAT I (L-methionine). The present data suggest that DL-homocysteine biphasically modulates KYNA synthesis. This seems to result from conversion of compound to S-adenosylhomocysteine, also acting dually on KYNA formation, and in part from the direct interaction of homocysteine with metabotropic glutamate receptors and KYNA biosynthetic enzymes. It seems probable that hyperhomocystemia-associated brain dysfunction is mediated partially by changes in brain KYNA level.
Protein kinase A and protein kinase C are involved in processes that enhance glutamate release at glutamatergic nerve terminals. However, it is not known whether these two kinases co-exist within the same nerve terminal, nor is it clear what impact their simultaneous activation may have on neurotransmitter release. In cerebrocortical nerve terminals, co-application of forskolin, which increases cAMP levels and activates protein kinase A, and 4beta-phorbol dibutyrate, a direct activator of protein kinase C, synergistically enhanced the spontaneous release of glutamate. This enhancement exhibited both tetrodotoxin-sensitive and tetrodotoxin-resistant components. Interestingly, the tetrodotoxin-resistant component of release was not observed when cyclic AMP-dependent protein kinase (PKA) and calcium- and phospholipid-dependent protein kinase (PKC) were activated separately, but developed slowly after the co-activation of the two kinases, accounting for 50% of the facilitated release. This release component was dependent on voltage-dependent Ca2+ channels that opened spontaneously after PKA and PKC activation and occurred in the absence of Na+ channel firing. These data provide functional evidence for the co-existence of PKA- and PKC-signalling pathways in a subpopulation of glutamatergic nerve terminals.
Extracellular signals may regulate mitogen-activated protein kinase (MAPK) cascades through a receptor-mediated mechanism. As a signaling superhighway to the nucleus, active Ras-MAPK cascades phosphorylate transcription factors and facilitate gene expression. In cultured rat striatal neurons, the present work systemically examined the linkage between glutamate receptors and the extracellular signal-regulated kinase 1/2 (ERK1/2) subclass of MAPK. We found that glutamate induced a rapid and transient phosphorylation of ERK1/2. Similar responses of ERK1/2 phosphorylation were also induced by the ligands selective for each of three subtypes of ionotropic receptors (NMDA, AMPA and kainate), although not by the subgroup-selective agonists for three subgroups of metabotropic glutamate receptors after 8-9 days in culture. The ERK1/2 phosphorylation induced by all ionotropic receptor agents was dose-, time- and Ca(2+) influx-dependent and occurred in neurons, but not glia. The NMDA-, AMPA- and kainate-induced ERK1/2 phosphorylation was blocked only by the antagonists selective for respective subtypes. The ERK1/2 phosphorylation induced by these agents was also sensitive to the MAPK kinase 1 (MEK1) inhibitor PD98059 and the MEK1/2 inhibitor U0126. In a further attempt to evaluate the role of active ERK1/2 in activating a downstream transcription factor cAMP response element-binding protein (CREB), NMDA, AMPA, and kainate were found to increase CREB phosphorylation. The NMDA- and AMPA/kainate-induced CREB phosphorylation was completely and partially blocked by U0126, respectively. These results revealed a positive linkage between ionotropic glutamate receptors and MEK-sensitive ERK1/2 phosphorylation in striatal neurons. The active ERK1/2 cascade activates the downstream transcription factor CREB to participate in the regulation of gene expression.
Hyperhomocysteinaemia has recently been recognized as a risk factor of cardiovascular disease. However, the action mechanisms of homocysteine (Hcy) are not well understood. Given that Hcy may be involved in the recruitment of monocytes and neutrophils to the vascular wall, we have investigated the role of Hcy in essential functions of human neutrophils. We show that Hcy increased superoxide anion (O2*-) release by neutrophils to the extracellular medium, and that this effect was inhibited by superoxide dismutase and diphenyleneiodonium (DPI), an inhibitor of NADPH oxidase activity. The enzyme from rat peritoneal macrophages displayed a similar response. These effects were accompanied by a time-dependent increased translocation of p47phox and p67phox subunits of NADPH oxidase to the plasma membrane. We also show that Hcy increased intracellular H2O2 production by neutrophils, that Hcy enhanced the activation and phosphorylation of mitogen-activated protein kinases (MAPKs), specifically p38-MAPK and ERK1/2, and that the migration of neutrophils was increased by Hcy. Present results are the first evidence that Hcy enhances the oxidative stress of neutrophils, and underscore the potential role of phagocytic cells in vascular wall injury through O2*- release in hyperhomocysteinaemia conditions.
CREB and Elk-1 phosphorylation by metabotropic glutamate receptors in striatal neurons
- Choe
- Es
- Wang
- Jq
Choe ES, Wang JQ (2002) CREB and Elk-1 phosphorylation by metabotropic glutamate receptors in striatal neurons. Int J Mol Med 9:3–10.
Role of group I metabo-tropic glutamate receptors and NMDA receptors in
- E Zieminska
- A Stafiej
- Lazarewicz
- Jw
Zieminska E, Stafiej A, Lazarewicz JW (2003) Role of group I metabo-tropic glutamate receptors and NMDA receptors in
Hyperhomocysteinaemia; with reference to its neuroradiological aspects
- Van den Berg
Mice deficient in cystathionine beta-synthase
- Watanabe
Glutamate-induced homocysteic acid release from astrocytes
- Benz
CREB and Elk-1 phosphorylation by metabotropic glutamate receptors in striatal neurons
- Choe
Folate deprivation induces neurodegeneration
- Ho